Food Intake Review 2005

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REVIEW

Brain regulation of food intake and appetite: molecules and

networks

C . B R O B E R G E RFrom the Department of Neuroscience, Karolinska Institute, Stockholm, Sweden

Abstract. Broberger C (Karolinska Institute,

Stockholm, Sweden). Brain regulation of foodintake and appetite: molecules and networks

(Review). J Intern Med2005; 258: 301327.

In the clinic, obesity and anorexia constitute

prevalent problems whose manifestations are

encountered in virtually every field of medicine.

However, as the command centre for regulating food

intake and energy metabolism is located in the

brain, the basic neuroscientist sees in the same

disorders malfunctions of a model network for how

integration of diverse sensory inputs leads to a

coordinated behavioural, endocrine and autonomicresponse. The two approaches are not mutually

exclusive; rather, much can be gained by combining

both perspectives to understand the pathophysiology

of over- and underweight. The present review

summarizes recent advances in this field including

the characterization of peripheral metabolic signals

to the brain such as leptin, insulin, peptide YY,

ghrelin and lipid mediators as well as the vagusnerve; signalling of the metabolic sensors in the

brainstem and hypothalamus via, e.g. neuropeptide

Y and melanocortin peptides; integration and

coordination of brain-mediated responses to

nutritional challenges; the organization of food

intake in simple model organisms; the mechanisms

underlying food reward and processing of the

sensory and metabolic properties of food in the

cerebral cortex; and the development of the central

metabolic system, as well as its pathological

regulation in cancer and infections. Finally,

recent findings on the genetics of human obesityare summarized, as well as the potential for novel

treatments of body weight disorders.

Keywords: brainstem, cerebral cortex, feeding,

hypothalamus, metabolic, reward.

Introduction

Few tasks executed by the brain hold greater

survival value than keeping us fed and in adequate

nutritional state. It is not surprising then that the

central nervous system has developed a meticu-

lously interconnected circuitry to meet this chal-lenge. A consequence of this organization is that an

energy-dense environment favours the development

of obesity, whilst overcompensation may shut down

the drive to feed. In todays society where evolving

disease demographics and lifestyle allow for a

greater diversity of metabolic phenotypes than

perhaps ever before [1] disorders of both extremes

of energy intake are common in health care.

This paper builds partly on presentations made at a

Nobel Conference on Brain Control of Feeding Behaviour

organized at the Karolinska Institute, Stockholm, Sweden,

in September 911, 2004.

Journal of Internal Medicine 2005; 258: 301327 doi:10.1111/j.1365-2796.2005.01553.x

2005 Blackwell Publishing Ltd 301


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Obesity is increasing at an alarming rate in

industrialized and developing countries alike [2]

and is associated with a wealth of conditions

afflicting virtually all organ systems [3, 4]. Exam-

ples diverge widely to include cholelithiasis [5],

osteoarthritis [6], infertility [7], stroke [8], cutane-ous infections [9], wound healing deficiencies [10],

as well as a general increase in mortality [11]

and social and professional stigmatization [12].

The urgency of the problem is illustrated dramat-

ically by the previous rarity of paediatric obesity-

associated type 2 diabetes, which is increasing to

the point of taking over as the leading cause of

childhood diabetes [13]. The opposite extreme of

anorexia and hypophagia includes not only anor-

exia nervosa [14] but is also a common and

potentially fatal complication of infections [15],

malignancies [16] and ageing [17].Unlike many other common diseases, these disor-

ders have an obvious solution: adjusting food intake

and exercise until normal body weight has been

restored. However, it is no great revelation that this

solution is as simple as it has repeatedly proved

elusive [18]. Experimental studies confirm the com-

mon knowledge that weight loss almost always is

followed by a rapid return to initial weight once the

anorexigenic regimen is terminated [19]. Notably,

the same applies to humans subjected to voluntary

overfeeding [20], supporting the concept of a tightly

regulated set-point for body weight. Treatment ofeating disorders has been remarkably unsuccessful

a consequence possibly of that we are battling

ancient systems maintained by thrifty genes that

favour the preservation of energy stores [21]. Avail-

able options for pharmacological therapy leave much

to be desired, and compounds that have been

introduced for obesity management have subse-

quently often been withdrawn due to intolerable

side-effects [22]. The most effective obesity treatment

at present is bariatric surgery, but this is a

complicated procedure not without adverse effects

[23]. Preventive measures have frustratingly lim-ited effect. It has proved even more difficult to

devise strategies for increasing food intake in cases

of anorexia. Although some success has been

reported with behavioural approaches for anorexia

nervosa [24], the more common forms of cancer-

and inflammation-associated anorexia remain a

major therapeutic challenge. Novel treatments are

greatly needed.

But what systems should such treatments target?

Early clinical observations that patients with pituit-

ary tumours and accompanying injury to the base of

the brain develop obesity [2528], inspired experi-

mental lesion studies [2933], which demonstrated

that damage to particular regions of the hypotha-lamus and brainstem lead to profound, often fatal,

alterations of feeding behaviour. It also became

apparent that signals from the peripheral energy

stores [34] and gastrointestinal canal [35] provide

essential cues for appetite and satiety. Based on

these and other findings, Stellar [36] half a century

ago proposed a dual centre hypothesis for the

initiation of motivated behaviour. The hypothesis

included both mechanisms for sensing peripheral

cues, separate nuclei (i.e. the dual centres) for

stimulating and inhibiting behaviour, and connec-

tions between the hypothalamus and higher brainregions to allow for internal states to determine the

threshold for initiating behaviour. Of all motivated

behaviours, the model is perhaps most applicable for

food intake. Yet, for all its heuristic value, the dual

centre hypothesis was as low on specifics as it was

laden with theory. Research dating in particular

from the last decade has changed this. Today, we

have an understanding of the circuitry and neuro-

pharmacology of feeding behaviour that can be

probed for therapeutic targets. The present article is

not an exhaustive review of the central control of

energy metabolism [37, 38], but summarizes recentadvances, which have brought new insight into the

peripheral signals describing the metabolic state to

the brain; the input stations in the hypothalamus

and brainstem sensitive to these signals, the organ-

ization of feeding behaviour in simple and complex

organisms; the link between food intake and energy

expenditure; the neural framework for integrating

metabolic cues and reward properties; the mecha-

nisms of infection- and cancer-associated anorexia;

developmental and genetic causes of obesity and

novel therapeutic strategies.

A central framework for sensing andorchestrating energy metabolism

The regulation of energy metabolism presents a

prototypical homeostatic system, with the brain

acting as the central coordinator and rectifier

(Fig. 1). It is one of the great wonders of the brain

that body weight stays remarkably fixed (as a

2005 Blackwell Publishing Ltd Journal of Internal Medicine258: 301327

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set-point) most of the time in most people [39]. The

first step in maintaining this homeostasis is for

the brain to inform itself of the metabolic status of

the individual, which it does through two main

channels. First, hormonal signals reflecting the

availability and demand for metabolic fuel is relayed

via neurones in the hypothalamus. The receptors for

these signals are primarily expressed on two neuro-

chemically distinct sets of neurones located in thearcuate nucleus (Arc) in the mediobasal hypothala-

mus, alongside the third ventricle [40]. One neurone

group expresses neuropeptide Y (NPY); increasing

NPY release or activation of these neurones by a

variety of approaches results in increased food

intake and decreased energy expenditure. The other

group expresses the neuropeptide precursor pro-

opiomelanocortin (POMC), which is processed to

melanocortin peptides; activation of these neurones

has the opposite effect of triggering the NPY cells, i.e.

decreased food intake and increased energy expen-

diture. The yinyang relationship between the twoArc groups is further underscored by their opposite

regulation by leptin and insulin, hormones signal-

ling metabolic affluence, which decrease the expres-

sion of NPY, whilst they increase the expression of

POMC. The second main input for information

pertaining to energy balance is the brainstem,

classically viewed as a channel for visceroceptive

information as cranial nerves, in particular the

vagus nerve, carrying information from the aliment-

ary organs enter the brain here. Vagal afferents

synapse onto [41, 42] and excite [43] neurones in

the brainstem nucleus tractus solitarii (nTS). Fromboth the hypothalamus and the brainstem, projec-

tions fan further into the brain to engage other brain

regions in the initiation and organization of food

intake. As in all homeostatic systems, the brain has

at its disposal three effector pathways to activate

when the controlled variable (i.e. body weight)

needs to be adjusted: behaviour (i.e. food intake), the

endocrine system and the autonomic nervous sys-

tem [44]. Importantly, all three of these systems are

engaged downstream of the Arc and nTS stations to

provide a synchronized response to fluctuations in

energy balance; the first primarily in the volitionalcontrol of intake, the latter two regulate energy

expenditure.

Peripheral control of feeding behaviour

Metabolic state is reflected in a diverse array of

signals of the brain. Recent investigations have shed

light on some of the key hormones and the vagal

ARC

POMC

NPY

GHRELIN

OEA

PYY

VAGUS

LEPTIN

DMX

IML

INSULIN

nTS

Fig. 1 The central metabolic circuitry is regulated by numerous

endocrine and neural inputs. Schematic illustration of how brain

networks regulating ingestive behaviour communicate with per-

ipheral organs. Hormones supplying information about the per-

ipheral metabolic state to the brain include the gastrointestinalpeptides ghrelin and PYY(3-36), insulin from the pancreas and

leptin from adipose tissue. Ghrelin and leptin act both on the

hypothalamus (Arc) and the brainstem (nTS). The afferent por-

tion of the vagus nerve innervates most of the gastrointestinal

tract where it collects information about the immediate aliment-

ary state, and terminates in the nTS. The lipid mediator OEA is

produced in the duodenum and activates the brainstem, possibly

via the vagus nerve. Both the Arc (via antagonistic NPY- and

POMC-expressing cells) and the nTS project further into the brain

in parallel pathways to engage higher brain regions into ingestive

behaviour. Outputs from the brain regulating energy expenditure

include both branches of the autonomic nervous system; the

sympathetic system whose preganglionic neurones are located in

the intermediolateral cell column (IML), which is directly inner-

vated by POMC neurones from the Arc, as well as the parasym-pathetic system with preganglionic neurones for the efferent

portion of the vagus nerve located in the dorsal motor nucleus of

the vagus (DMX). The efferent autonomic innervation regulates,

e.g. glucose homeostasis via actions in liver and skeletal muscle.

See text for details.

2005 Blackwell Publishing Ltd Journal of Internal Medicine 258: 301327

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mechanisms that shape the central response to

nutritional challenges (Fig. 1).

The vagus nerve

The gastrointestinal canal is equipped with a myriadof sensory receptors along its full crown-rump

extension [45]. Thus, the taste, texture and

mechanic stress of food is reported to the brain to

provide an online description of the immediate

alimentary state. This information is routed to the

nTS primarily via the afferent portion of the vagus

nerve, so that vagal activation causes satiation and

meal termination. (Parallel neural feedback is also

supplied by sensory neurones innervating the oral

cavity mediating, e.g. taste, and lesser-studied

splanchnic nerves [46].) Vagal afferents are broadly

sensitive to gastrointestinal signals, including gas-troduodenal distension, the presence of chemically

distinct nutrients within the gastrointestinal tract as

well as peptides produced by endocrine cells in the

gut wall, most prominently cholecystokinin (CCK), a

well-characterized satiety signal [47, 48]. Import-

antly, these signals are integrated within the indi-

vidual vagal sensory neurone prior to the signal

being relayed in the nTS [49, 50]. The neurochem-

ical identity of viscerosensory vagal neurones has

remained enigmatic, but these cells likely signal via

glutamate [51] and the neuropeptide cocaine- and

amphetamine-regulated transcript [52], whichinhibits feeding upon brainstem administration [53].

Leptin

An appetite-regulating endocrine signal from fat

tissue maintaining energy homeostasis had been

hypothesized already with parabiosis experiments

in the 1950s [34]. The seminal discovery of leptin,

the adipocyte-derived protein hormone providing

this signal, by Friedman and collaborators in 1994

[54] was a decisive catalyst for much of the current

investigation on peripheral modulation of centralnetworks. A little more than a decade later, leptin

has been shown to modulate several aspects of

energy balance through several different mecha-

nisms and across a wide spectrum of timeframes,

alerting the brain to the state of body adiposity [55]

and acting as a fat-o-stat. It is now well estab-

lished that the pronounced obesity in genetically

leptin-deficient ob/ob mice is due to the loss of a

centrally active feeding-inhibitory messenger, asrestitution of the leptin signal in these animals

normalizes food intake and body weight [56]

(Fig. 2). Serum leptin correlates well to the size of

the body fat deposit, and falls with weight loss [57].

This relationship is seen also in obesity, where the

combination of hyperleptinaemia and hyperphagia

has led to the suggestion that overweight is

characterized by leptin resistance [58]. Central

actions underlie both leptin-mediated feeding sup-

pression as well as the extensive peripheral meta-

bolic effects of this hormone; thus, e.g. replacement

of leptin receptors selectively in the brain of ob/obmice is sufficient to prevent hepatic steatosis [59].

Insulin

Insulin is well recognized as the key glucostatic

regulator.Recent data demonstrate that in addition to

the control of peripheral glucose uptake, this role also

encompasses powerful central effects, in synergism

with leptin. First, intracerebroventricular (i.c.v.)

administration of insulin decreases food intake [60]

via insulin receptors expressed on Arc neurones [61].

The role of insulin in feeding is complicatedby thefactthat the hypoglycaemia resulting from elevations in

serum insulin in itself stimulates food intake. How-

ever, when blood glucose changes are compensated

for, hypophagia is seen also with increases in periph-

eral insulin [62, 63], suggesting that the central

effects of insulin are anabolic. (This secondary hypo-

glycaemia may also explain why the insulin secretion

triggered already at the sight of a palatable-looking

Fig. 2 Genetically leptin-deficient ob/ob mice treated subcutane-

ously (s.c.) with saline (left) or with leptin (right). The severe

obesity in these animals is abolished with leptin replacement

therapy. Figure generously provided by Dr Jeffrey M. Friedman.




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meal stimulates appetite (the cephalic phase [64]),

an indication that direct sensory input relayed via the

cortex can set off powerful appetitive mechanisms.)

Secondly, and again similar to leptin, insulin also

modulates peripheral energy metabolism via central

effects by inhibiting liver gluconeogenesis [65, 66].Thus, whilst the brain does not depend on insulin for

glucose uptake, it is very much interested in what

insulin has to say about the metabolic state of the

body.

Peptide YY (3-36)

In addition to CCK, several gut-derived peptides

provide alimentary feedback to the central metabolic

circuits [67]. Peptide tyrosine-tyrosine (3-36)

[PYY(3-36)], a member of the NPY peptide family

produced by enteroendocrine cells [68], has recentlybeen shown to act as an important feedback signal

from the gut to the hypothalamus. Following a

meal, PYY(3-36) is released into the circulation

[69], specifically stimulated by the presence of lipids

and carbohydrates in the lumen of the distal ileum

and colon [70, 71]. Peripheral administration of this

hormone inhibits food intake and causes weight loss

[72, 73]. While some laboratories initially were

unable to replicate this effect [74], this may partly be

due to discrepancies in technique [75] and the

results have since been independently confirmed

[76, 77]. The satiety effect of PYY(3-36) is compar-atively modest but, importantly, is observed also in

humans, including obese patients [73, 78]. Plasma

levels of PYY(3-36) rise markedly following ileal

resection [79, 80], an observation that has been

linked to the weight loss observed in patients

undergoing this procedure (S.R. Bloom and C. Le

Roux, personal communication).

Ghrelin

Thus, the gastrointestinal-brain axis has long been

viewed as a key channel subserving meal termin-ation with CCK and PYY(3-36) as prime mediator

examples. Novel findings on the hormone ghrelin

(produced in stomach and small intestine epithelia

[81], see [82]) are challenging this doctrine. Ghrelin

is unique as the first gut hormone shown to

stimulate food intake [67]. Both peripheral and

central injections of ghrelin result in increased

feeding as well as fat mass [8386]. Ghrelin levels

peak sharply in anticipation of a meal in humans as

well as experimental animals [87], resulting in

stimulation of both feeding and gastric emptying

[88] through actions possibly involving the vagus

nerve [89], and may thus provide a meal initiation

signal. In the hypothalamus, peripherally adminis-tered ghrelin mainly activates the NPY neurones

[85, 90] and antagonizing the actions of these cells

inhibits the orexigenic effect of ghrelin administra-

tion [85, 91]. In contrast, the melanocortin pathway

does not appear to be primarily involved [90]. Recent

reports have proposed that ghrelin is synthesized also

in hypothalamic neurones, but this issue remains

controversial, in part due to the contradictory claims

of the site of central ghrelinergic neurones and the

failure to demonstrate ghrelin mRNA in the brain (cf.

[92] and [93]).

Importantly, a loss of the hunger message relayedby ghrelin has been suggested as the mechanism

behind the weight-reducing effects of bariatric sur-

gery [94]. The initial rationale for gastric bypass

[95] was that the procedure would produce weight

loss by means of malabsorption. However, this turns

out to be a transient effect due to the considerable

compensatory potential of the digestive system.

Nevertheless, weight loss persists, caused instead

by a loss of appetite and hypophagia [96]. Concom-

itant with this effect, a fall in plasma ghrelin is

observed following the bypass procedure, in contrast

to the ghrelin increase associated with nonsurgicalweight reduction, where weight relapse is common

[86, 97]. (Note, however, findings that argue

against such a relationship, see [67].) Furthermore,

clinical data tie the hyperphagia observed in Prader

Willi syndrome to strikingly high plasma ghrelin

levels [98]. These results, coupled with the discovery

that elevated plasma ghrelin is a marker for future

weight gain (D.E. Cummings and J. Krakoff, personal

communication) indicate that interfering with ghre-

lin signalling offers a clinically promising approach

to treating eating disorders.

Oleoylethanolamide

A role for endogenous cannabinoids in appetite

regulation has long been suspected from the carbo-

hydrate craving observed in marijuana smoking

[99]; indeed, increased appetite is a diagnostic

criterion for cannabis intoxication [100]. Neuronal

production of cannabinoids is widespread and these




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mediators play an important and general role in the

modulation of synaptic transmission [101], with the

orexigenic effects likely mediated via central cann-

abinoid CB1 receptors [102, 103]. However, the

lipid family to which the cannabinoids belong also

includes other members with opposite and periph-eral effects on energy metabolism. Piomelli and

colleagues have accumulated evidence that the fatty

acid oleoylethanolamide (OEA), chemically but not

pharmacologically similar to the cannabinoids, is

produced in the duodenum and acts via the vagus

nerve to decrease body weight through activation of

the nTS [104]. OEA increases the inter-meal latency,

an effect distinct from that of, e.g. CCK, which

primarily decreases meal size [105]. However, chan-

ges in energy expenditure also underlie the OEA-

mediated weight reduction and are especially pro-

nounced in models of obesity, involving in particularincreased fat utilization, whereas glucose home-

ostasis is relatively unaffected [106]. The catabolic

effects are most noticeable as a slowing of body

weight gain in growing rats, with OEA synthesis

reduced by food deprivation and stimulated in

response to increased demands on energy availability

such as cold exposure [104]. The metabolic actions

of OEA depend selectively and critically on genomic

as well as nongenomic actions of the ubiquitous

nuclear peroxisome proliferator-activated receptor-

alpha (PPAR-a) [107]. These results add obesity to

the growing list of potential therapeutic applicationsfor nuclear receptor pharmaceuticals. Notably, drugs

that target PPAR-a, e.g. gemfibrozil, are already in

clinical use to treat hypercholesterolaemia [108].

Integration of peripheral signals in the Arc

The peripheral signals described above thus act upon

the Arc (and nTS, see below) to influence the central

pathways regulating energy balance. In the Arc,

receptors for leptin and insulin found on NPY and

POMC neurones serve to inhibit transcription of NPY

[109, 55] and increase POMC mRNA levels [110112] via differential second messenger systems [113].

It is becoming evident that insulin, leptin and other

metabolically relevant hormones eventually con-

verge not only on a common set of neurones, but

indeed also on the same molecules. Recent reports

highlight the role of the ATP-dependent potassium

current, IK(ATP), as a molecular target mediating

rapid, electrophysiological effects, of peripheral

hormones. This K+ conductance is a priori sensitive

to the availability of metabolic fuel as a fall in

intracellular levels of the energy donor ATP causes

the channel to open, leading to K+ influx and

hyperpolarization; this mechanism enables neurones

expressing IK(ATP) to vary their excitability in responseto changes in glucose concentration[114].Leptin and

insulin both hyperpolarize Arc neurones by enhan-

cing IK(ATP) [115, 116], by activating a common

enzyme, phosphoinositide 3 (PI3) kinase [116, 117].

It should be emphasized that the transmitter pheno-

type of Arc neurones expressing IK(ATP) is a contro-

versial issue which remains to be conclusively

resolved [118120]. Additional signals likely weigh

in on IK(ATP); this current is augmented when the

concentration of fatty acid derivatives is increased

locally within the Arc by inhibition of lipid oxidation,

a message of energy surplus that also decreases foodintake [66, 121]. This convergence of nutrient

information makes the PI3-kinase/IK(ATP) a key

integration node within the metabolic signalling

chain, attractive as a therapeutic target.

Modulation of the membrane potential of Arc

neurones has recently been demonstrated to control

glucose homeostasis. Opening of Arc K(ATP) chan-

nels via either hyperinsulinaemia or central inhibi-

tion of lipid oxidation inhibits vagal efferent (i.e.

parasympathetic) gluconeogenic signals to the liver,

promoting the use of fat as metabolic fuel [66, 122].

The Arc is also the site of central leptin regulation ofglucose homeostasis as selective restoration of Arc

leptin receptor expression in otherwise leptin recep-

tor-deficient mice is sufficient to correct their

hyperglycaemia [123]. These results show that

insulin modulates glucose homeostasis by independ-

ent peripheral and central mechanisms and empha-

size that interconnectivity within brain metabolic

regions serve to switch the body between different

fuel sources, in parallel to controlling food intake.

Interestingly, in obese rats, hypothalamic IK(ATP)channels fail to respond to leptin and insulin [115,

116]. Whether similar defects underlie insulin and/or leptin resistance in human diabetes and obesity is

an interesting possibility, which remains to be

investigated.

Output from the Arc

NPY neurones. Neuropeptide Y is one of the most

potent stimulators of feeding known [124], an




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effect that has been confirmed by various approa-

ches [40]. While there is conflicting data on whe-

ther deletion of the NPY gene produces hypophagia

(cf. [125] and [126]), the obesity of ob/ob mice is

attenuated when combined with an NPY)/) geno-

type [127], suggesting that NPY is an importantmediator of central leptin signalling. Stimulation of

feeding appears to be transduced predominantly via

postsynaptic NPY Y1 receptors, as determined from

pharmacological and genetic engineering studies

(reviewed in [128], Fig. 3a). However, the syner-

gistic actions of multiple NPY receptor subtypes

participate to produce orexigenic effects in vivo

[129]. Detailed behavioural analysis of those effects

suggests that NPY primarily stimulates appetitive

rather than consummatory behaviour [130].

POMC neurones. Pro-opiomelanocortin is a large

precursor protein which gives rise to several bioac-

tive peptides. Among these, the melanocortin pep-tides, in particular a- and c-melanocyte-stimulating

hormone, have been shown to exert potent ano-

rexigenic effects when administered i.c.v. [131,

132]. Central melanocortin effects are mediated by

the melanocortin 3 and 4 receptors (MC3R and

MC4R, respectively; Fig. 3b). Deletion of the genes

for either POMC, MC3R or MC4R result in obesity in

mice, suggesting that the melanocortin system is

crucial in maintaining body weight [133135] as

supported by similar findings in humans (see below).

MC4R)/) mice also increase their feeding in

response to a high fat diet, in contrast to wild-typelittermates where a reduction is seen andob/obmice,

which maintain the same intake as with regular

chow [136], underlining the importance of the

melanocortin system for adjusting food intake in

response to caloric variations. In addition, the hall-

mark hypophagia seen in disease models as diverse

as renal failure, immunological challenge with

lipopolysaccharide (LPS) and tumour implants is

ablated. The obesity in MC4R-deficient animals is

partly due to changes in energy expenditure, such as

deficient diet-induced thermogenesis [136]. The

anatomical substrate for this effect may be a directprojection from the POMC neurones in the Arc to

the preganglionic sympathetic neurones in the spi-

nal cord [137139] constituting a link between the

metabolic integrator and the autonomic effector

system. Interestingly, the spinal projection sets the

POMC neurones apart from the neighbouring NPY

neurones which otherwise exhibit very parallel

innervation patterns. However, it should be pointed

out, that in humans, the melanocortin system

appears to be more geared towards regulating feed-

ing behaviour, with a proportionately smaller role in

peripheral metabolism [140].

NPYPOMC interactions. Interactions between the

Arc populations allow the NPY neurones to control

the activity of the POMC cells via two mechanisms.

First, NPY neurones coexpress agouti gene-related

peptide (AGRP), an endogenous melanocortin ant-

agonist [141143]. Thus, at the axon terminal,

melanocortin action can be blocked by simultaneous

Fig. 3 Expression of NPY and melanocortin receptors in the

mouse brain. In situ hybridization histochemistry (a) shows the

distribution of NPY Y1 receptor mRNA detected as silver grains in

a coronal section, revealing dense expression in the cerebral

cortex and nuclei in the amygdala, thalamus and hypothalamus.

In panel b, green fluorescent protein (GFP) is expressed in a

neurone under control of the melanocortin (MC) 4 receptor pro-

moter; note strong immunoreactivity throughout cell soma and

dendrites. A Nissl-stained coronal section (c) shows neurones

clustered to form the paraventricular hypothalamic nucleus

(PVH) alongside the third ventricle. The PVH constitutes a central

integrative hub within the metabolic circuitry. (d) Immunoh-

istochemical for GFP (indicating the presence of the MC4 receptor)

and in situ hybridization for NPY Y1 receptor mRNA have beencombined in a section from the amygdala, revealing the coexist-

ence of these receptors in neurones downstream of the Arc. Figure

produced by Drs Toshiro Kishi and Joel K. Elmquist. Reprinted

with permission from Macmillan Publishers Ltd.; Molecular Psy-

chiatry 2005;10:132146.




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release of AGRP, and in agreement with such an

arrangement, a single i.c.v. administration of AGRP

causes an impressively long-lasting (one week)

suppression of food intake [144]. Secondly, at the

cell body level, POMC neurones are innervated by

NPY-ergic terminals [145] and express the Y1receptor [146], through which NPY causes a

powerful membrane potential hyperpolarization (i.e.

inhibition) [147]. Surprisingly, no reciprocal inner-

vation has yet been described and Roseberry et al.

[147] did not detect any changes in the electrical

properties of NPY neurones using a melanocortin

analogue. Thus, there may exist an asymmetrical

interaction in the Arc favouring the orexigenic

NPY/AGRP message over anorexigenic melanocor-

tin signalling. However, an inhibitory influence over

the NPY neurones may be provided by PYY (3-36),

which is a selective agonist of the inhibitory Y2autoreceptors [148] expressed by these cells [146].

Such gastrointestinal negative feedback has been

proposed as the mechanism whereby PYY(3-36)

inhibits feeding as no such effect is observed in mice

genetically deficient for the Y2 receptor and appli-

cation of the peptide inhibits the electrical activity of

Arc NPY terminals [73]. This effect is relatively

selective as disruption of other relevant metabolic

pathways does not affect the satiety effect; the

PYY(3-36) effect persists both after vagotomy and in

MC4)/) mice [76], suggesting that neither the nTS

nor the Arc POMC neurones are directly involved.

Classical transmitters: glutamate and GABA. While

much of the current research on the central regula-

tion of energy balance focuses on the role of peptides,

it should be emphasized that in the hypothalamus, as

in the rest of the brain, the key chemical mode of

communication between neurones is via amino acid

transmitters, i.e. excitatory glutamate and inhibitory

c-amino butyric acid (GABA). Indeed, in the absence

of glutamate and GABA-mediated transmission, little

remains of hypothalamic synaptic activity [149,

150]. The major function of peptides, in addition totheir genomic effects, is likely to modulate the syn-

aptic transmission of classical transmitters [151]

with which they coexist [152]. Interestingly, glu-

tamate N-methyl-d-aspartate (NMDA) receptors

have been found to stimulate feeding with remark-

able anatomical specificity within the lateral hy-

pothalamic area (LHA), in comparison with other

hypothalamic regions tested and the amygdala

[153]. Infusion of NMDA antagonists locally within

the LHA blocks both agonist-induced and depriva-

tion-induced food intake, indicating the involvement

of endogenous glutamatergic tone in natural feeding

[154]. Histochemical studies suggest that within the

Arc, NPY neurones largely contain GABA, whereasPOMC neurones signal via glutamate [155, 156].

Downstream targets of the Arc. The downstream

cellular effects of NPY are still mysterious. It was

initially assumed that feeding-promoting neurones

in loci sensitive to NPY orexigenesis were excited by

NPY. However, all known members of the NPY

receptor family couple to inhibitory second mes-

senger systems [128]. Electrical excitation has been

proposed to come about in the form of disinhibition

via NPY-mediated suppression of GABA-dependent

inhibitory postsynaptic currents [157, 158], withmelanocortin stimulation producing the opposite

result, i.e. inhibition via stimulation of GABA release

[157]. However, that does not explain the role of

postsynaptic Y1 receptors, which exist throughout

the hypothalamus [146, 159, 160] (Fig. 3a). The

most potent orexigenic effects of NPY are seen

within the perifornical region/LHA [124], where

NPY/AGRP-ergic terminals appear to target two

separate populations of neurones expressing the

neuropeptides hypocretin (Hcrt; also known as

orexin) and melanin-concentrating hormone (MCH;

Fig. 4, [142, 161, 162]). This pathway is of interestas Hcrt and MCH potently modulate wakefulness

[163167], providing a means for metabolic signals

to control arousal state. Surprisingly, in a recent

investigation of the electrophysiology of the LHA

neurones, melanocortin stimulation did not affect

the electrical properties of MCH-expressing cells,

whereas both these and Hcrt-expressing cells were

inhibited by NPY [168, 169]. Furthermore, micro-

injection of NPY into the LHA appears to activate a

group of neurones distinct from those expressing

Hcrt or MCH [170]. As with all neural interactions,

it is important to bear in mind that the activity ofneurones can be influenced via several independent

mechanisms, including (but not exclusively) elec-

trical and transcriptional effects, and that different

changes proceed along different temporal scales.

Thus, a functional Arc-LHA pathway cannot be

excluded. Nevertheless, these data invite a re-eval-

uation of the role of Hcrt and MCH as downstream

mediators of the NPY and POMC neurones.




9/27

Circadian regulation of metabolic processes. In addition

to the various controls summarized above, metabolicprocesses also follow strict circadian variations as

recently underscored by the demonstration that

inactivation of key genes maintaining circadian

rhythmicity results in manifest metabolic syndrome

in mice [171]. Thus, for example, in rats the active

period of the day is immediately preceded by

coordinated peaks in hepatic glucose output (via the

sympathetic nervous system) and glucose uptake in

striated muscle (a parasympathetic effect; see [172]).

Buijs et al. [173] have investigated which brain re-gions are responsible for this synchronization. Using

anatomical tracing they find that the chains of

neurones innervating liver and muscle are separated

all the way through brainstem and hypothalamus to

distinct populations of preautonomic master neu-

rones in the suprachiasmatic nucleus [173], the

brain region maintaining circadian rhythmicity and

entrained by direct retinal input [174, 175]. The

CerebralCortex

AcbSh

PFCx

MCH Hcrt

LHA

Othersubcortical nuclei,

incl. BST,MPO, PVT,

DMH,Amgdl,Raphe, PAG

and PBN

NPY POMC

Arc

DMX

IML

FOODINTAKE

ENERGYEXPENDITURE

INGESTIVE(Motivated)BEHAVIOUR

Sympathetic

ParasympatheticANS

Endocrine

regulationincl. thyroid andadrenocortical axes

PVH

Pituitary

Fig. 4 Integration in higher brain regions determines the central response to changes in peripheral metabolic state. Schematic illustration

of connections between brain regions responsible for coordinating the behavioural somatomotor (i.e. food intake), autonomic and endocrine

(the latter two regulating energy expenditure) responses that together constitute the motivated ingestive behaviour used by the nervous

system to meet nutritional challenges. The antagonistic orexigenic NPY and anorexigenic POMC neurones in the Arc project in parallel

paths to numerous subcortical nuclei [including the bed nucleus of the stria terminals (BST), the medial preoptic area (MPO), the

paraventricular nucleus of the thalamus (PVT), several hypothalamic nuclei, e.g. the dorsomedial nucleus (DMH), the amygdala (Amgdl),

the serotonin-containing system in the raphe nuclei, the periacqueductal grey area (PAG) and the parabrachial nucleus (PBN)] distributedthroughout the brain. A projection to neurones expressing melanin-concentrating hormone (MCH) or hypocretin (Hcrt) in the lateral

hypothalamic area (LHA) provides an indirect pathway to the cerebral cortex for metabolic signals relayed via the Arc. The cortex in turn

projects back heavily to both the LHA and other feeding-regulatory regions. In addition, the LHA also receives an inhibitory input from the

shell of the nucleus accumbens (AcbSh), which in turn is modulated via prominent excitatory inputs from the prefrontal cortex (PFCx).

Thus, the LHA is positioned to integrate both homeostatic and reward-related signals in the gating of food intake. Energy expenditure is

modulated via outputs from the Arc to neuroendocrine neurones in the paraventricular hypothalamic nucleus (PVH), which control

release of, e.g. thyrotropin-releasing hormone and adrenocorticotropic hormone from the pituitary gland. Energy expenditure is also

regulated by projections from POMC neurones in the Arc and descending pathways from the PVH to autonomic preganglionic neurones in,

e.g. the dorsal motor nucleus of the vagus (DMX; parasympathetic) and spinal cord intermediolateral cell column (IML; sympathetic).

Note that ascending projections from the brainstem, which provide parallel important metabolic inputs to the brain, have not been included

in the figure. See text for details.




10/27

distinct pathways originating in the suprachias-

matic nucleus are particularly interesting in con-

junction with the discovery that the autonomic

inputs to intra-abdominal and subcutaneous fat

stores are also separate [176]. In humans, shift work

[177] and sleep deprivation [178] are associatedwith increased adiposity, findings that have been

linked to the sleep-associated peak in leptin secretion

[179]. However, this anatomically separate inner-

vation indicates that loss of periodicity in the circa-

dian input to adipose tissue may disrupt the balance

between different fat compartments leading, in turn,

to manifestations of the metabolic syndrome, which

is correlated to abdominal but not subcutaneous fat

accumulation [180].

Contributions of hind- and forebrain to

feeding regulation

As mentioned above, the brainstem provides a port

for vagal and other neural sensory signals into the

brain. Classical accounts of brain regulation of

feeding described two systems balancing each other:

the hypothalamus, monitoring the periphery for

signals alerting central circuits to diminishing

energy stores, and the brainstem, receiving oral

and gastrointestinal information as an online signal

of the amounts and qualities of the food that was

being ingested. This arrangement would allow the

hypothalamus to function as a long-term controlorchestrating meal initiation and the brainstem

served as a short-term control for meal termination.

Much of our knowledge on the different contribu-

tions of the fore- and hindbrain in meal regulation

comes from a lesion model developed by Grill and

Norgren [33]. Disconnecting the forebrain (which

includes the hypothalamus) produces a rat inca-

pable of the motor activation necessary for normal

feeding. However, if this animal whose brainstem

remains intact is provided sucrose solution via an

intraoral cannula, intake can be measured as the

solution consumed until the meal is terminated asthe animal lets solution drip out of the mouth. These

decerebrated rats maintain the ability to terminate

their meal in response to changes in gastrointestinal

feedback, but are unable to compensate for varia-

tions in the caloric value of the fed solution,

resulting in anorexia if the sucrose concentration

is reduced. Similarly, removal of the post-oral

feedback (by e.g. vagus nerve transection or gastric

drainage) leads to increases in meal size as well as

duration [181, 182], although there is a compen-

satory delay in the latency to meal initiation,

possibly mediated by the hypothalamus. These

results underscore the role of the Arc as a metabolic

sensor. It should be pointed out that an intact Arc isnot necessary for meal initiation humans and

animals with selective lesion of this region not only

eat, they eat copiously [29, 183185].

It is now becoming evident that the brainstem can

integrate much the same signals as have been

shown to modulate hypothalamic activity. Leptin

receptors are expressed at several strategically

located brainstem sites, and selective stimulation of

these receptors suppresses food intake at doses

comparable with those used in forebrain injections

[186, 187]. Here, leptin activates the same medial

region of the nTS that is stimulated by gastricdistension [188], suggesting an anatomical site of

integration of long- and short-term feeding controls.

Likewise, melanocortin agonists can reduce feeding

and body weight by brainstem mechanisms [189].

Interestingly, the neurones in the nTS mediating the

viscerosensory signal may also be POMC-encoded

[43, 190], a finding that puts our understanding of

melanocortin-mediated meal suppression in a new

light. Orexigenic effects of ghrelin are also seen with

selective local administration both in the hypotha-

lamus [84] and in the brainstem [191] (Fig. 5a,b).

Finally, glucosensitive neurones have been recordedin the nTS [192]. Thus, the nTS is in no way a

Fig. 5 Ghrelin increases food intake following brainstem admin-

istration. Unilateral injection of 10 pmol (but not 5) of ghrelin

(black bars) into the dorsal vagal complex, including the nucleus

tractus solitarii, results in a significant increase of food intake both

1.5 and 3 h after drug administration compared with vehicle

(white bars), in an experiment by Faulconbridge et al. [191].

Reprinted with permission from the American Diabetes Associ-

ation;Diabetes 2003;52:22602265.




11/27

passive transducer of viscerosensory signals, but

serves to integrate numerous indices of the animals

metabolic state. This conclusion is supported by the

observation that the hyperphagia of rats that lack

leptin receptors is caused by larger meal size (i.e.

meal termination delay) rather than an increasednumber of feeding bouts [193], suggesting an action

localized in the hindbrain. However, the response to

CCK (i.e. brainstem satiety signalling) as well as

normal meal size is restored in these animals

following selective re-establishment of leptin recep-

tor expression in the Arc. Weighed together, these

data emphasize that actions of metabolically rele-

vant hormone take place at a few, but distributed,

sites in the brain contributing to a coordinated

feeding response.

Beyond the primary sensors: CNSintegration of hunger and satiety signals

As in all matters involving the brain, behaviour begs

the question: What are the pathways? For the

metabolic signals to produce behaviour they need to

proceed further into the brain beyond the primary

sensors in the Arc and nTS and ultimately engage

regions that initiate and organize behavioural,

autonomic and endocrine response patterns. Histo-

chemical studies have revealed that (i) the Arc

projections diverge widely throughout the brain

[194197] including, via indirect pathways, amassive cortical innervation [198] (Fig. 4), (ii) the

NPY and POMC populations project in remarkably

parallel paths [196] and may converge on the same

cells as supported by the widespread coexistence of

Y1 and MC4 receptors [199] (Fig. 3d), and (iii) the

nTS innervates largely the same nuclei as the

ascending projections from the Arc [196, 200

202]. This circuitry indicates that integration

between the primary metabolic sensors in the Arc

and nTS is a distributed phenomenon, and it has

been suggested that this arrangement allows for

motivational state to be weighed into the networkbefore reconvergence and the final decision for a

proper metabolic response is made [40]. In addition

to this scheme, extensive reciprocal projections

connect the Arc with its target regions [203]. The

functional implications of this arrangement are not

clear at present. There may exist a sequential

arrangement hidden within the network that has

so far eluded the techniques used to investigate the

system. Another alternative is that the reciprocity

may produce a reverberating signal, such as the

large-scale oscillations described in, e.g. thalamo-

cortical systems [204]. Possibly, such persistent

activity may be important in the triggering and

maintenance of an anabolic response. The hierar-chical organization of the metabolic circuits and its

relationship to behaviour presents a major future

scientific challenge. (For further discussion of the

systems organization of energy balance regulation,

the reader is referred to recent exhaustive reviews

[37, 38].)

Coordinating the metabolic output

It also remains to explain how the divergence

convergence organization of the Arc/nTS projec-

tions interdigitates with the efferent networksunderlying the final metabolic response. As

already mentioned, motivated behaviours (classic-

ally divided into ingestive, reproductive and defen-

sive) involve three distinct outlets: components of

the autonomic and neuroendocrine systems as

well as coordinating the overall behaviour of the

animal [44]. Activity within these three effector

systems is hypothesized to be organized by a

collection of cell groups within the medial hypo-

thalamus area, collectively termed the hypotha-

lamic visceromotor pattern generator network

[205], which subsequently recruits elements with-in a behavioural control column, spanning the

mes- and diencephalic midline [206]. Separate

groups of control column nuclei produce ingestive,

reproductive and defensive behaviours, but con-

nections between these networks allow them to

interact for purposes of mutual exclusion so that

only one behaviour is expressed at once [207].

The paraventricular nucleus (PVH; Fig. 3c) is a

crucial control column module for ingestive beha-

viour. The PVH collects metabolic information

from oropharyngeo- and viscerosensory receptors

and humoral signals (both directly and via theArc), and is regulated by biological rhythms via

the suprachiasmatic nucleus and by the overall

state of the animal as reported from the cerebral

hemispheres via relays in the septum [206].

Output from the PVH, in turn, employs all

three outlets described above: endocrine via neuro-

endocrine neurones controlling pituitary hormone

release, autonomic via direct projections to




12/27

preganglionic neurones in the spinal cord inter-

mediolateral cell column (sympathetic) and the

dorsal motor nucleus of the vagus (parasympathe-

tic), and behavioural (i.e. somatomotor) via several

pathways innervating the brainstem [137, 208

210] (Figs 1 and 4). The hierarchical arrangementupstream of these motor nuclei bears some simi-

larity to the basal ganglia organization for the

control of conscious movement [206]. The details

of the underlying anatomy remain mysterious, but

it is clear that, rather than a simple sequential

organization where information flows neatly from

one collection of neurones to another, we are

faced with an interconnected series of hubs that

collect, integrate and disseminate information.

Food intake in lower organisms: models for

the organization of behaviour

Intriguingly, an improved understanding of the

organization of feeding behaviour is now coming

from studies of lower organisms. Animals such as

the nematode Caenorhabditis elegans and the sea slug

Aplysia present several technical advantages: beha-

viour is easily divided into discrete sequences, the

nervous system consists of a limited number of

neurones whose electrical properties and intercon-

nectivity has been characterized in detail, and, in the

case ofC. elegans, the genome is highly accessible for

molecular manipulation. This knowledge makes itpossible to understand how nature organizes phys-

ical networks to efficiently initiate, organize and

terminate behaviour.

Caenorhabditis elegans

Feeding in C. elegans is polymorphic; depending on

genetic background animals will feed either alone or

in aggregates [211213]. Naturally occurring var-

iations in a single amino acid position of the

neuropeptide receptor NPR-1 (for neuropeptide

receptor resemblance-1) translate into either asolitary or a social feeding phenotype [212]. Acti-

vation of NPR-1, by shifting network properties,

leads to activation of social feeding behaviour, and

the amino acid substitution in NPR-1 determines the

response to stimulation with the neuropeptide

ligands flp 18 or )21 [214]. Conversely, null

mutations in the npr-1 locus alter the balance in

favour of solitary feeding [214]. Thus, a single gene

is sufficient to redirect behaviour. Intriguingly, the

predicted transmembrane domains of NPR-1 display

considerable homology to mammalian NPY recep-

tors [212], suggesting that similar molecular com-

ponents underlie related behaviours throughout

evolution. Recent data demonstrate that within thisnetwork an important upstream regulator of NPR-1-

mediated feeding is oxygen concentration, showing

how external cues reset behaviour [215, 216].

Aplysia

In Aplysia, a meticulously dissected network has

been highly informative in characterizing the neural

mechanisms underlying various behavioural com-

ponents. Aplysia feeding can be initiated by sensory

stimulation of the lip and is consolidated by arousal

caused by the exposure to food. In parallel with thefeeding central pattern generator (CPG) circuit, a

network controlling arousal is triggered, which then

feeds back into the CPG [217]. Termination of food

intake is achieved by switching ingestion to the

opposite behaviour of egestion [218]. Separate

motor neurones within the CPG effectuate ingestion

and egestion. Switching the balance between these

neurones is elegantly accomplished by recruiting a

single additional interneurone into the circuitry via

an electrical synapse [219]. Feeding patterns in this

social mollusc also differ substantially in the absence

or presence of other animals; company is animportant incentive for feeding. Pheromones secre-

ted from other Aplysia stimulate a neurone directly

driving the appetitive phase of feeding, which also

excites a control neurone for the consummatory

phase so that these behaviours are properly organ-

ized, leading to larger and more frequent eating

bouts [220]. These studies provide information on

how organization within a neural system translates

into behaviour, with potentially broad implications

considering the surprisingly large overlaps between

human and mollusc feeding behaviour. Importantly,

they may shed light on the process of selectionwithin the behavioural repertoire and why we eat in

apparent violation of homeostatic mechanisms

when our fat stores are filled to the brim.

Food reward: role of the nucleus accumbens

However, to elucidate the neural organization of

feeding in complex vertebrates, it is necessary to




13/27

consider that our choice of food is not simply a

function of energy supply and demand, but also very

much linked to reward value. Homeostatic systems

active within the brain operate at the mercy of

motivational states. The motivational state is a

product of, inter alia, limbic influences relayed inpart via the amygdala, and reward factors. Obvi-

ously, adding reward experience to food is a means

of, e.g. avoiding consumption of foods whose taste

indicate the presence of invasive microorganisms,

but also promoting those whose taste signals

particular nutritional value. As a result, pairing

feeding with pleasure may override normal satiety

mechanisms, resulting in hyperphagia and obesity.

The concept of reward is intimately linked to that of

addiction, and it has been suggested that obesity is a

consequence of an addiction to food [221]. Drugs of

abuse converge upon the mesolimbocortical systemto produce reward, specifically by enhancing dop-

amine release in the nucleus accumbens (Acb) of the

forebrain as a final common pathway [222]. There

is little doubt that changes in dopaminergic trans-

mission affects food intake. Indeed, animals unable

to produce brain dopamine die of starvation unless

fed by gavage [223], and a common side-effect of

neuroleptics affecting dopamine signalling is obesity

[224]. However, novel data suggest that dopamine

primarily acts to reinforce behaviours at the initial

encounter with a novel reward, but the release of

dopamine decreases once the behaviour has beenestablished within the behavioural repertoire [225].

Thus, whilst dopamine modulates learning and

locomotion associated with motivational behaviour,

it appears not to modulate feeding behaviour per se;

blocking Acb dopamine signalling does not alter

total food consumption in starved rats, although it

suppresses ambulation associated with feeding

[226].

However, transmitter systems other than the

dopaminergic system connect Acb to components

of the metabolic circuitry [227]. Notably, chronic

access to a preferred flavour (chocolate-fat solution)produces the same transmitter changes in the Acb

as chronic morphine or ethanol, suggesting a

common reward mechanism for palatable food

and conventional drugs of abuse [228]. These

changes include increased transcription of opioid

peptides such as enkephalin. In turn, stimulating l-

opioid receptors in the Acb increases intake of fat-

enriched foods with high palatability value [229],

which may be interpreted as positive reinforcement.

This effect is not seen following inhibition of neural

transmission in the basolateral amygdala (BLA) or

the LHA [230]. The connection between the BLA

and forebrain cortical regions has been implicated

in gauging food palatability, and an intact BLA isrequired for determining the reward value embed-

ded in sensory input [231]. In contrast, the central

amygdala (CeA) serves more like a general gate-

keeper of feeding as inactivation of this subnucleus

blocks consumption of all foods [230]. This dichot-

omy may be explained by the different projections

of the BLA (innervating higher forebrain regions

such as the prefrontal cortex) and the CeA (aimed

towards the postulated hypothalamic and brain-

stem feeding pattern generators). The connection

between the Acb and the LHA has also been shown

to modulate food intake (Fig. 4). Kelley andcolleagues have shown that blocking excitation of

the GABA-encoded Acb results in hyperphagia, an

effect contingent upon intact transmission in the

LHA [232]. Glutamatergic excitation in the Acb is

predominantly supplied by the cortex, so that this

cortex-Acb-LHA pathway could offer the prefrontal

cortex in particular a channel for inhibiting feeding

behaviour in favour of other behaviours [227].

These data indicate that in the regulation of

ingestive behaviour, the Acb is indeed at the

interface between motivation and action as ori-

ginally postulated for this structure [233].

Studies on the food intake experience inhumans and monkeys: relevance forunderstanding obesity

Defining the cortical involvement in feeding beha-

viour is likely vital for understanding human

eating disorders. Crosstalk between the cerebral

cortex and primary sensors is extensive; impres-

sively, the hypothalamus provides the largest

external cortical input with the exception of the

thalamus [198], and reciprocal connections areplenteous [234]. Investigations of monkeys and

humans by Rolls and collaborators have revealed

that the sensory properties of food are processed in

two steps in the cortex [see 235]. The insular

cortex, functionally and neuroanatomically impli-

cated as viscerosensory cortex [236], acts as a

primary taste cortex where these individual fea-

tures, i.e. taste, appearance, smell and texture, are




14/27

represented to determine which food is being

ingested. Individual neurones are sensitive to

single stimuli, and do not adapt their firing even

when the stimulus (e.g. glucose) has been present

for a long time [237]. The orbitofrontal cortex

(which receives direct input from the insularcortex), however, acts as a higher-order taste

cortex, which determines how pleasant a particular

food is. Orbitofrontal neurones are broadly tuned

to react to multiple sensory features, so that the

firing of these cells result from the combined

inputs from several sensory modalities [238],

although different cells respond to different quan-

titative combinations of stimuli. The subjective

pleasantness rating is proportional to orbitofrontal

activation and this activation drops accordingly

when a particular food is eaten to satiety [239]

(Fig. 6). Thus, computations within the orbitofron-tal cortex confer hedonic qualities upon the

feeding circuitry, a role whose importance in

human appetite may be underestimated when

extrapolating from rodent studies. The orbitofron-

tal cortex feeds directly into the LHA, which may

thus constitute an important nexus for linking the

subjective experience of food with homeostatic

signals.

The relationship of the hedonic/pleasure experi-

ence to human obesity has begun to be addressed in

neuroimaging studies, further emphasizing the cor-

tical parcellation of different feeding-related process-

ing. Whereas during hunger, activation is observed

predominantly in regions associated with the regu-

lation of emotions (limbic and paralimbic cortex),

satiety is followed by activation in the prefrontalcortex, postulated to play a role in the inhibition of

inappropriate behaviours [240]. The presentation of

a palatable sweet solution after a day and a half

of fast resulted in increased signal from the insular

cortex [241]. Moreover, almost all observed changes

are accentuated in obese compared with lean

subjects, i.e. both increases and decreases in activity

are of greater amplitude [241] (Fig. 7). These

differences are not exclusively accounted for by the

hyperglycaemia and hyperinsulinaemia manifest in

obese subjects. The functional implication of the

intensified patterns of activation and inactivation inobese subjects is not clear at present, but may be of

pathophysiological importance. Indeed, these re-

sponses persist also after weight loss in postobese

subjects [242] suggesting that they are not a

consequence of the overweight as such, although

it is not known if such accentuated responses are

seen also prior to the development of obesity.

Elucidating these mechanisms may be significant

also for understanding the human response to food

advertisements.

10

20

40

0 50 80

+1

+2

0

-1

-2

pe

on

ct

banana odour

mango odour

30

mL

Volume of 20% blackcurrant juice

food

satiating

response to

Behavioural

Firing rate

(spikes s1)Blackcurrant odour

Spontaneous

Fig. 6 Neurones in the orbitofron-

tal cortex decrease their firing in

response to a particular food as that

food is consumed to satiety. Extra-

cellular recording from an odour-

responsive neurone in the orbito-

frontal cortex of a male macaque.

The firing rate of one neurone in

response to different odours was

recorded whilst the animal con-

sumed a blackcurrant juice. As

satiety increases (lower panel), the

electrophysiological response toblackcurrant odour diminishes,

whereas the response to unrelated

odours [banana, mango, phenyl

ethanal (pe), citral (ct), onion (on)]

is unaffected or elevated. Figure

produced by H.D. Critchley and

E.T. Rolls and is used with permis-

sion from the American Physiolo-

gical Society; J Neurophysiol

1996;75:16731686.




15/27

Development of the feeding circuitry

A series of recent studies have shed light both on the

normal ontogenetic development of hypothalamic

circuits as well as how these processes are influenced

by the nutritional state of the young animal, with

important implications for the aetiology of obesity. At

birth, the hypothalamus is rather sparsely inner-

vated by Arc NPYergic fibres, and in, e.g. the PVH asubstantial NPY/AGRP innervation is seen first at

postnatal day (P) 15 [197, 243, 244]. Similarly,

whilst Arc NPY and AGRP mRNAs are detectable

from birth, levels peak at P15, and drop to adult

levels by P30 [244] (Fig. 8). This development

parallels the maturation of the ability to regulate

suckling in response to caloric demands [245],

linking changes within the Arc metabolic sensor

and adult control of feeding. Notably, failure in the

development of the Arc is associated with fatal

anorexia in a genetic model [246, 247]. It should be

mentioned, that during the early postnatal period,

and under certain physiological circumstances, tran-

sient expression of NPY in other hypothalamic nuclei

can be seen [243, 248]. The role of these transitory

NPY projections remains to be determined.

As the Arc provides a main conduit for leptin, it isperhaps not surprising, given these data, that pups

are unable to respond to leptin by changing their

food intake [249], nor to ghrelin [250]. Yet, there is

a pronounced peak in serum leptin prior to weaning

[251]. Recent data from Bouret et al. [252] suggest

that a postnatal leptin surge is essential for the

development of Arc projections. In adult ob/obmice,

there is a distinct paucity of Arc-derived terminals.

Fig. 7 Lean and obese subjects show differences in brain activation during different states of hunger. Statistical parametric maps of

significant brain responses (P 0.005, not corrected for multiple comparisons) to hunger and early satiety in obese (top row) and lean

(bottom row) subjects, respectively, at 4 mm above (left images), 4 mm below (middle images), and 16 mm below (right images) a

horizontal plane passing through the anterior and posterior commissures (coordinates from the Montreal Neurological Institute). The right

hemisphere in each section is on the readers right. The T-value colour-coded areas were regions of the brain in which significant

changes in blood flow (a marker of neural activity) were detected in response to hunger (from yellow to white, in increasing order of

T-value), as stimulated by a 36-h fast, or in response to early satiety (from blue to green, in increasing order of T-value), as stimulated by

consumption of a satiating liquid meal [304]. The figure is intended for visual inspection only of several brain regions, where significantly

greater responses in obese compared with lean individuals were detected, including the middle temporal gyrus (TEMP) insula (INS),dorsolateral prefrontal cortex (DLPFC), hippocampus (HIPP), temporal pole (T.POLE), orbitofrontal cortex (OFC), and ventrolateral

prefrontal cortex (VLPFC). Figure generously provided by Dr Angelo DelParigi.




16/27

However, administration of leptin to ob/ob mice

during early development, but not in adulthood,

results in innervation patterns similar to what is

seen in lean littermates, in parallel with a normal-

ization of body weight. These data provide a novelmechanism for how nutritional signals in the early

postnatal stage exerts long-lasting effects on the

metabolic wiring in the adult. It will be of interest to

determine if similar mechanisms are at play in the

development of the brainstem-vagal system. In this

context, the importance of the prenatal metabolic

environment should also be remembered. Gesta-

tional diabetes and obesity is associated with obesity

in the offspring of both rats [253] and humans

[254]. An extensive long-term study is currently in

progress to investigate what changes can be seen in

central metabolic circuits in nonhuman primates

following intrauterine exposure to diabetes [255].Conversely, changes in hypothalamic circuitry dur-

ing senescence may contribute to the loss of appetite

that often accompanies ageing [17]. Such changes

are observed in older rats, who display significantly

lower levels of expression of NPY [256] and POMC,

as well as POMC-positive neurones [257] and

diminished dendritic arbours [258] in the arcuate

nucleus compared with younger animals.

Fig. 8 Arc NPY/AGRP projections to the PVH develop during the postnatal period in the rat. Figure displays confocal micrographs of

double-label immunofluorescence for NPY (red) and AGRP (green) in the PVH. Double-labelled fibres are shown in yellow. These images

demonstrate that at postnatal day 5 (P5) that there are minimal Arc NPY/AGRP projections to the PVH, whilst there is an abundance

of NPY fibres that originate from other sources. By P10 there is a significant concentration of Arc NPY/AGRP projection in the PVH, but

they do not reach the adult levels until around P15. Images represent a 10- lm thick collection of optical sections collected at 0.5-lm

intervals. Images were captured with a 25 oil objective (0.75 NA) and represent an area of 400 400 lm. Figure generously provided

by Dr Kevin Grove.




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Mechanisms of anorexia in infection andcancer

The central systems mediate not only obesity but also

anorexia of various aetiologies. Infection-based anor-

exia is a well-established and clinically highly rele-vant model [259], which has been instrumental in

uncovering the pathways through which microor-

ganisms cause reduced food intake. Bacterial compo-

nents such as LPS from Gram-negative bacteria and

muramyl dipeptide from Gram-positive bacteria acti-

vate CD14- and Toll-like receptors on host T-cells to

induce production of cytokines such as interferon-c;

these results are corroborated by experiments

employing genetic removal of strategic components

along the pathway [260]. Cytokines are produced

peripherally, but activate vagal afferents [261]. In

addition, cytokines stimulate prostaglandin produc-tion via the enzyme cyclooxygenase 2 (COX-2) in

cerebral endothelial and perivascular cells [262]. This

latter pathway is important for the induction of LPS-

induced anorexia, and can be blocked with indo-

methacin and other, more selective, COX-2 inhibitors

[263]. Further downstream, the prostaglandins acti-

vate neurones producing serotonin (5-HT; [264]), a

known anorexigenic transmitter [265, 266], which

in turn has recently been shown, via 5-HT2Crecep-

tors, to stimulate melanocortin signalling [267].

Thus, the signalling cascade set off by pathogenic

bacteria ultimately results in activation of the centralanorexigenic system. These results may shed light

also on the anorexia accompanying noninfectious

inflammatory conditions. In this context it is inter-

esting to note that, based on structure and signalling

pathways, leptin itself belongs to the cytokine family

[268, 269].

While anorexia is an important component also of

wasting in cancer patients, nutritional supplements

only alleviate part of the cachectic syndrome [270],

which accounts for a fifth of cancer deaths [271].

Cachexia differs from starvation in that both adipose

and lean mass is lost, whereas starvation primarilydecreases fat stores [272]. A main explanation for

this relationship is that many cancer tumours

secrete proteins, e.g. proteolysis-inducing factor

[273], which suppress protein synthesis in skeletal

muscle, via activation of an ubiquitin proteolytic

pathway [274]. Parenteral nutrition may thus be of

very limited value to the patient if such catabolic

mechanisms are not interrupted. However, it turns

out that a polyunsaturated fatty acid found in fish,

eicosapentaenoic acid (EPA), suppresses the activity

of the proteolytic complex whilst simultaneously

inhibiting tumour growth [275]. This finding has

promising bedside implications; adding EPA to the

diet of cancer patients attenuates muscle degrada-tion and stabilizes body weight [276].

From rodent to human: genetic dissection

In few fields of medicine is the question of nature

versus nurture more immediate than in the regula-

tion of body weight. While it is evident that the

incidence of obesity has accelerated far more rapidly

than can be explained by population shifts within the

genome, it is also true that individual differences

determine how we react in an energy-dense environ-

ment. Aptly summed up by Olden and Wilson [277]:genes load the gun, environment pulls the trigger.

Studies of monozygotic twins show that the heritable

component of obesity equals that of height and

surpasses virtually every other major disease studied,

e.g. breast cancer, schizophrenia, cardiovascular

disease [278280]; some 40% of obesity can be

attributed to genetic causes [281]. In a series of

elegant studies, in particular by ORahilly, Farooqi

and their colleagues, it has been shown that muta-

tions in several components of the anorexigenicsignal

chain, including leptin [282] (Fig. 9), the leptin

receptor [283], POMC [284] and neuropeptide pro-cessing enzymes [285] result in severe early-onset

obesity in humans. While these monogenic disorders

incontrovertibly demonstrate that genetic abnormal-

ities can cause obesity, it cannot automatically be

concluded that mutations and sequence variants are

common causes of the metabolic syndrome.

However, it is now becoming apparent that

mutation of a particular key signalling protein, the

MC4R [286], accounts for as many as 5% of cases

of severe obesity [140, 287289]. There is a

remarkably solid relationship between the severity

of the mutation, as revealed in in vitro assays, andthe size of a test meal consumed by the patient

[140]. Indeed, the heritable component is almost

exclusively represented by increased intake of

energy, with only a small component accounted

for by changes in resting metabolic rate [140]. (In

contrast, in the mouse MC4R)/) counterpart,

deficient energy expenditure is an important factor

underlying overweight [136].) The sometimes




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modest phenotypes observed in rodent gene knock-

out experiments (e.g. NPY gene deletion [125])

have been interpreted as evidence for a high

degree of redundancy within the system under-

lying metabolic regulation. However, genetic stud-

ies such as these (as well as, notably, observed in

nematodes [212]), suggest that there are weakpoints distributed throughout the system where

minute changes in nucleotide sequence can have

profound effects on the expression of behaviour.

These vulnerable links along the metabolic signal-

ling chain offer promising targets for therapeutic

intervention.

Therapeutic prospects

As pointed out in the Introduction, effective treat-

ments for eating disorders are short in supply and

urgently needed. While much can be gained simplyby increased efforts to educate patients in the

benefits of weight loss and exercise [290, 291], it

is also clear that such therapies in many cases fail in

the absence of adequate pharmacological buttress.

The anti-obesity drugs presently used in clinical

practice have relatively modest effects, and others

still have been withdrawn due to intolerable cardio-

vascular adverse effects [22]. These drugs have often

targeted broadly distributed systems, e.g. serotonin

pharmacology, and it is perhaps not surprising that

alterations ensue within multiple body functions.

Another issue to consider is the timing of adminis-

tration of feeding-regulatory drugs, which may be

more crucial than in any other therapeutic applica-tion (T. Bartfai, personal communication). However,

based largely on the research summarized above,

several novel compounds are now being tested in

clinical trials, phases II and III [22, 292]. Many of

these compounds target neuropeptide systems. The

selective neuroanatomical distribution of many

neuropeptides may provide an advantage in

attempting to minimize side-effects, and whilst the

field of peptide-based pharmaceuticals has not lived

up to initial hopes, the clinical experience from

opioid drugs such as morphine and naloxone [293]

suggest that interfering with peptide signalling canproduce powerful neuropsychiatric effects in

humans [294]. In particular, drug development

initiatives have centred on the melanocortin system

[295], encouraged by the human genetic

data reviewed above, and several nonpeptide com-

pounds are now being tested in a clinical setting

[22].

The cannabinoid system also provides an attract-

ive target, with promising early results with the

inverse CB1 receptor agonist, rimonabant [296],

currently in phase III clinical trial. Initial evalua-

tions suggest that rimonabant decreases bodyweight by ca 510%, with beneficial effects also on

insulin resistance and dyslipidaemia [292]. Interest-

ingly, this drug has the added benefit of facilitating

smoke cessation. However, the full safety profile of

rimonabant has not yet been released. Conversely,

the active component of marijuana, D9-tetrahydro-

cannabinol, is also being evaluated for treatment of

AIDS-associated anorexia [297].

Early hopes for a leptin-based obesity regimen

were quelled by clinical trials showing only very

limited weight loss following leptin administration to

obese subjects [298], and there are also indicationsthat such treatment may in itself produce the leptin

resistance hypothesized to be a part of obesity [299].

It should be pointed out, however, that in the rare

cases of genetic leptin-deficiency, treatment with

recombinant leptin has virtually normalized body

weight in near-fatally obese children, whilst simul-

taneously inducing age-appropriate puberty [300]

(Fig. 9). The same treatment has also been used to

Fig. 9 Leptin deficiency in humans responds to leptin treatment.

A 3-year-old boy with congenital leptin deficiency with severe

obesity (body weight 38 kg; BMI SD 7.2) (left). On the right,

the same patient, after four years of daily subcutaneous admin-istration of recombinant leptin. Leptin treatment results in a

dramatic decrease in adiposity (body weight 29 kg; BMI SD

0.9) and normalization of all metabolic abnormalities including

hyperinsulinaemia. Figure generously provided by Drs Sadaf

Farooqi and Stephen ORahilly.




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successfully rectify several of the metabolic abnor-

malities associated with lipodystrophy [301, 302].

Furthermore, pertaining to a not uncommon diag-

nosis, leptin treatment has recently been demon-

strated as a highly promising therapy for functional

amenorrhoea [303].

Conclusion

In summary, significant advances in our under-

standing of feeding behaviour have been achieved

using a combination of clinical, behavioural, elec-

trophysiological, anatomical, genetic and imaging

techniques. This investigation has defined an under-

lying circuitry and neuropharmacology, which can

now be probed for therapeutic targets. Thus, whilst

many questions remain to be answered, in partic-

ular regarding the causal mechanisms of obesity andanorexia as well as the central circuitry bridging

metabolic input and output, there is reason for hope

in offering effective treatments for these exception-

ally common and debilitating disorders.

Conflict of interest statement

No conflict of interest was declared.

Acknowledgements

The author gratefully acknowledges generous finan-cial support from Wenner-Gren Stiftelserna, Vet-

enskapsradet, Jeanssons Stiftelser, Hagbergs Stiftelse,

Rut och Arvid Wolffs Stiftelse, Thurings Stiftelse,

Svenska Lakaresallskapet, Ake Wibergs Stiftelse, Kgl.

Vetenskapsakademien, Hedlunds Stiftelse, Magnus

Bergvalls Stiftelse, Lars Hiertas Minne, Axel Linders

Stiftelse, Langmanska Kulturfonden, Teodor Neran-

ders Fond and internal funds of Karolinska Institutet.

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