Hormones and the control of porphyrin biosynthesis and structure in the hamster Harderian gland

MICROSCOPY RESEARCH AND TECHNIQUE 34:123-132 (1996)

Hormones and the Control of Porphyrin Biosynthesis and Structure in the Hamster Harderian Gland A.P. PAYNE, S.W. SHAH, F.A. MARFt, J. McGADEY, G.G. THOMPSON, AND M.R. MOORE Departments of Anatomy (AS’?., S.W.S.. F.A.M., J.M.) and Medicine and Therapeutics (G.G.T., M.R.M.), Glasgow University, Glasgow G12 8QQ, Scotland

KEY WORDS Harderian gland, Hamster, Porphyrin

ABSTRACT The hamster Harderian gland seems to present both an excellent model for the control of porphyrin biosynthesis and an unusually robust example of the interrelationship between structure and function. It has been known for some time that 1) the capacity for manufacturing and storing porphyrins and 2) gland histology and ultrastructure are controlled by androgens. Thus, in intact males as well as in gonadectomised animals of either sex treated with androgens, porphyrin synthesis by the Harderian gland is suppressed and the gland tubules characteristically possess two cell types, the cytoplasm of both containing polytubular complexes. By contrast, the Harderian glands of intact females and castrated males synthesise and store large amounts of protoporphyrin, while their tubules possess only one cell type which lacks polytubular complexes. So overarching is the effect of androgens that they have been described as a “coarse tuning” effect on the gland. By contrast, the role of the ovary is both less dramatic and less well understood. In female hamsters, ovariectomy leads to degenerative changes in Harderian gland tubules and (probably) a release of stored porphyrin; a t the same time there is a reduction in enzyme levels and new synthesis. The causative hormone in this “fine tuning” is unclear at present. There is now clear evidence that the Harderian gland is also controlled directly by pituitary hormones. In particular, the use of continuous infusion osmotic minipumps has allowed us to demonstrate not only 1) that the expected rise in porphyrins and feminisation of gland morphology does not occur in castrated males receiving the dopamine agonist bromocriptine, but that 2) the simultaneous administration of prolactin does permit these changes; furthermore, 3) the administration of prolactin alone increases porphyrin synthesis above the levels found in untreated castrates. Similarly, bromocriptine administration to ovariedomised females markedly reduces porphyrin synthesis and masculinises gland structure; again, this is reversed by the simultaneous administration of prolactin. Prolactin must therefore be seen as equipotent with androgens in determining gland structure and activity. 8 1996 Wiley-Liss, Inc.

INTRODUCTION In a recent review, Payne (1994) highlighted many

basic gaps in our understanding of the Harderian gland, including 1) whether the gland is purely exo- crine or if it has an endocrine role also, 2) why, in one large, widespread, and successful group of mammals- the rodents-the gland manufactures and stores pho- totransducing molecules, porphyrins, and 3) why the gland appears to be controlled by so many factors. It is the latter characteristic which forms the basis for this paper.

In one species alone, the golden hamster, there is evidence that gland structure and biosynthetic activity are controlled by androgens, by the ovary, by thyroid hormones, and even directly by pituitary products such as gonadotrophins and prolactin. In this multiplicity, are we simply unable to distinguish between important and less important factors, or do all of them have real biological significance? Do they control the same aspect of gland function or a variety of different ones?

This paper seeks to review the comparative effects of the testis, the ovary, and prolactin on gland structure and activity in one species, the golden hamster.

HORMONE RECEPTORS Androgen receptors occur in the Harderian glands of

the male rat (Gustafsson and Pousette, 1975) and both sexes of the golden hamster (Vilchis and Perez-Pala- cios, 1989; Vilchis et al., 1987). Vilchis et al. (1992) reported that the androgen receptor in the Harderian glands of rats, guinea pigs, and mice shared common binding characteristics with that in the hamster; they also reported androgen receptors in ducks, chickens, marine turtles, and lizards. The widespread occurrence of androgen receptors throughout the Harderian glands of terrestrial vertebrates suggests a well-con- served intracellular macromolecule. In amphibia, an androgen receptor occurs in the Harderian gland of both sexes of the green frog R a m esculenta (d’Istria et al., 1991).

Receptors are also present for other steroid hor-

k i v e d January 1.1995; accepted in revised form March 16,1995. Address reprint requests to Dr. A.P. Payne, Department of Anatomy, Univer-

sity of Glasgow, Glaagow G12 SQQ, Scotland, UK.

0 1996 WILEY-LISS, INC,

124 A.P. PAYNE ET AL.

mones. For example, receptors for oestradiol have been found in the Harderian glands of the female armadillo (Weaker et al., 1983) and golden hamster (Vilchis and Perez-Palacios, 1989); Vilchis et al. (1992) also reported oestrogen receptors in the glands of rats, mice, guinea pigs, and rabbits (especially the pink lobe), while Vilchis et al. (1991) found oestrogen receptors in the glands of ducks and chickens. Progesterone receptors have not been found in mammalian Harderian glands, but may occur in the glands of reptiles (Vilchis et al., 1992), ducks, and chickens (Vilchis et al., 1991). Finally, there may be glucocorticoid receptors in the Harderian glands of ducks (Butler et al., 19781, hamsters (Vilchis and Perez-Palacios, 1989), and other rodents (Vilchis et al., 1992).

EFFECTS OF GONADAL HORMONES ON GLAND STRUCTURE AND ACTIVITY

Much of our knowledge in this area has come from studies using the golden hamster, a species in which the Harderian gland exhibits several sex differences (sum- marised in Payne, 1994; Payne et al., 1992a) (Fig. 1).

1. The presence of two epithelial cell types in the male gland, distinguished on the basis of lipid vacuole size. Type I cells possess numerous small lipid vacuoles, while type I1 cells possess fewer, larger vacuoles. Only one cell type occurs in the female gland, possess- ing relatively small vacuoles.

2. The presence in male gland cells of polytubular complexes, believed to derive from vacuolated smooth endoplasmic reticulum (Johnston et al., 1987). These are absent from the female. By contrast, the female gland cells contain membranous structures arranged in lamellae which are not present in large numbers in the male (Bucana and Nadakavukaren, 1972).

3. Interstitial mast cells, which are some 40 times more numerous in female Harderian glands than in male glands (Payne et al., 1982).

4. Levels of porphyrin and the activity of porphyrinogenic enzymes which are higher in female than in male glands (Lin and Nadakavukaren, 1982; Thomp- son et al., 1984). There may also be sex differences in the proportions of different porphyrins formed (Spike et al., 1990).

There is a clear hormonal basis to these sex differences, since castration of the adult male will convert all gland characteristics to the female pattern within a few weeks (see Table 1, together with Lin and Nadaka- vukaren, 1979; Payne et al., 1977; Woolley and Worley, 1954), a change which can be prevented by androgen replacement (Hoffman, 1971; Payne et al., 1977). Sim- ilarly, administration of androgens to adult females will result in the appearance of type I1 cells and polytubular complexes and a marked decrease in both porphyrin content and porphyrinogenic enzyme activity (Spike et al., 1985; Sun and Nadakavukaren, 1980). This remarkable interconversion suggests that the two cell types in the male gland are not independent, since type I1 cells appear de novo in the female gland after androgen administration and can be restored, following loss, in the glands of castrated males. This suggests they are two forms of the same cell, perhaps denoting

different activity or secretory states (Payne et al., 1992b). There are some other reported sex differences, including the proportions of C16 and C18 fatty acids within the gland (Lin and Nadakavukaren, 19811, protein composition (Hoh et al., 1984), metallic ion concentrations (Hoffman and Jones, 1981), indoles (Hoff- man et al., 1985; Menedez-Pelaez et al., 1988, 1989) and somatostatin (Puig-Domingo et al., 19881, and some of these are also altered by androgen manipula- tion.

The role of the ovary in controlling Harderian gland structure and activity in the female hamster is less dramatic (Spike et al., 1985,1986a). Ovariectomy leads to progressive degenerative changes within the gland (shown in Fig. 2), during which the epithelium becomes grossly attenuated and disappears in places so that porphyrin stores which were previously intraluminal be- come relocated within the interstitium. These abnor- mally located stores are usually surrounded by foreign body giant cells. During epithelial degeneration, neu- trophils invade the gland lumen in large numbers and present a highly vacuolated appearance. Latterly, in- dividual macrophages are encountered within the interstitium containing porphyrin spicules. Porphyrino- genic enzyme activity becomes progressively reduced, but the porphyrin content of the gland remains re- markably constant. This suggests that porphyrin stores are neither being lost from the Harderian gland nor being added to by means of new synthesis. Similar changes occur in aged, postreproductive female hamsters (Spike et al., 1988). There is no evidence of masculinisation of gland structure or activity following ovariectomy, except where this is combined with androgen administration (see above). It is not clear which ovarian product normally acts on the gland, though the gland possesses receptors for oestradiol (Vilchis and Perez-Palacios , 1989).

The effects of androgens (or their absence) on the hamster Harderian gland are so pervasive that it has been referred to a “coarse tuning,” determining whether the gland has a male or female morphology and whether it synthesises porphyrins or not. By contrast, the effects of the ovary on the female gland have been likened to a “fine tuning” (Payne, 1990, 19941, perhaps maintaining gland structure and determining gland productivity in different reproductive phases.

The golden hamster is the only mammalian species so far examined in which there is such a marked sexual dimorphism of Harderian gland structure. However, there is a general trend for the Harderian glands of female rodents to contain more porphyrin than the glands of males-for example, in the mouse (Shirama et al., 19811, the gerbil (Meriones unguiculatus) (Johnston et al., 1983), and the plains mouse (Pseudo- mys australis) (Johnston et al., 1985). The hormonal basis of this sex difference has received little attention. Shirama et al. (1981) found that androgens decreased the porphyrin content of the Harderian glands of female C3H/He mice, while Ulrich et al. (1974) found no effect of androgens on the porphyrin content of the Harderian gland in ovariectomised rats.

Hormones may also affect the nonmammalian Hard- erian gland. Thus, seasonal changes which occur in the

HORMONAL CONTROL OF HAMSTER HARDERIAN GLAND 125

Fig. 1. A tubule from the Harderian gland of (a) a normal male and (b) a normal female hamster. The male gland exhibits two cell types, distinguished by the size of their lipid vacuoles: type I (I) cells with numeroue small vacuoles and type I1 (II) with a few large ones. The cytoplasm of both cells contains numerous polytubular complexes

(PTC). The female gland has tubules composed of only one cell type. The gland synthesises porphyrins which are stored intraluminally (POR). The interstitial tissue of the female gland is rich in mast cells (MC).

Harderian gland of the edible frog (Ram esculenta), such as epithelial height and secretion rate, are controlled by hormones (d'Istria et al., 1991). Recently,

Varriale et al. (1992) used in vitro studies to show 1) that total gland RNA showed seasonal maxima which coincided with peaks of androgen receptors, 2) that

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phyrin production by the Harderian gland and feminisation of its structure. However, Buzzell et al. (1989) discovered that the administration of the dopamine agonist bromocriptine prevented both the expected rise in porphyrin biosynthesis and morphological feminisation. This finding is extremely important, but it is unclear in this situation how or where bromocriptine is having its action. The most likely site of action would be the hypothalamo-pituitary axis, where dopaminergic neurons control several neuroendocrine systems (Andersson and Eneroth, 1987). Firstly, we must con- sider which neuroendocrine systems could be acted upon by bromocriptine.

Dopamine and Gonadotrophin Release (See Fig. 3a)

Neurons containing gonadotrophin releasing hormone (GnRH) occur in a variety of diencephalic regions including the preopticlseptal region, anterior hypothalamus, arcuate and suprachiasmatic nuclei, bed nucleus of the stria terminalis, and organum vasculosum of the lamina terminalis (for reviews and interspecies differences see Barraclough, 1992; Clarke, 1987). Dopamine appears to have an inhibitory effect on cells in the me- dial preoptic area which secrete luteinising hormone releasing hormone (LHRH) at terminals in the median eminence (Kuljis and Advis, 1989). At midcycle, sero- tonergic neurons either suppress these inhibitory dopaminergic cells or override their action by sensitising LHRH neurons to excitatory influences by noradren- ergic neurons. There are also influences on LHRH neurons by y-amino butyric acid (GABA) and opiates (Fig. 3a). As a dopamine agonist, bromocriptine could be suppressing LHRH release and, hence, the raised LH levels known to occur after castration through negative feedback mechanisms.

If this were the reason for the effects of bromocriptine on the Harderian gland, it could imply that the gland is acted upon directly by gonadotrophins. This would be unusual but not impossible. Recently, Menen- dez-Pelaez et al. (1992) have reported that human chorionic gonadotrophin can accelerate the increase in porphyrin concentrations (and the levels of mRNA for the rate-limiting enzyme S-aminolaevulinate-synthase) which occurs after castration in the male hamster; there does not seem to be a comparable effect on the female gland. The possibility that a gonadotrophin might act directly upon the Harderian gland was en- visaged by earlier authors such as Clabough and Nor- vell(1973). There is no evidence at present for suitable receptors within the gland.

Dopamine and Prolactin Release (See Fig. 3b) Dopamine can affect prolactin release €?om the pitu-

itary gland. Indeed, dopamine has long been recognised as Prolactin Inhibitory Factor (PIF) (Bedonathan, 1985; Foord et al., 1983; Lamberts and MacLeod, 1990; Swennen and Denef, 19821, although very low levels of dopamine have a paradoxical stimulatory effect on prolactin release (Denef et al., 1980). It is not wholly clear which factors normally augment prolactin release, although it is widely supposed that thyrotrophin releasing hormone (TRH) (Thomas et al., 1988) or vasoactive

a. t LH

TRH 7P4 @&/ LONG

PORTAL

PORTAL IACTOTROPHS PITUITARY

I- 3

INTERMEDIATE LOBE CELLS

OESTRADIOL VEIN PROLACTIN b.

TSH C.

Fig. 3. A diagrammatic representation of the control of (a) luteinising hormone (LH), (b) proladin, and (c) thyroid stimulating hormone (TSH) by the hypothalamo-pituitary axis. The diagrams are not intended to be exhaustive, but each demonstrates a dopaminergic influence (DA) on the control mechanism. A, adrenaline; GABA, y-amino butyric acid; LHRH, luteinising hormone releasing hormone; NA, noradrenaline, TRH, thyrutrophin releasing hormone; 5HT, se- rotonin. See text for details.


TABLE 2. The porphyrin content (nmollg tissue) and the activity of the rate-limiting enzyme for porphyrin synthesis 5-amino-laevulinate s y n t h e (MA-s ) (nmol ALA formedlhlg protein) in the Harderian glands of untreated male hamsters (Control), in males eastmted for 6 weeks (Cast), and in males castrated for 6 weeks and treated with bromocriptine (Cast +- bro), bromocriptine plus proloctin (Cast +

bro + om). or Drolactin alone (Cast + pro)’ ~

Porphyrin ALA-s Number content activity

Control 8 78 2 11* 331 f 69* Cast + bro 6 104 % 10* 405 2 83* Cast + bro + pro 5 408f55 * 2,108 f 861* Cast 8 128 f 73 * 2,059~ 22 * Cast + pro 4 1,355 2 255 * 2,5102 4 * F P < 0.001 P < 0.001

‘See text far details. All figures are meana + SEM. Groups are significantly different (P < 0.05) if their asterisks are not in vertical alignment.

intestinal peptide (VIP) (Akema et al., 1988; Kato et al., 1984) is involved. One complicating feature is that prolactin occurs in prerelease and release forms, and prolactin stores which are either less than 1 h or more than 12 h old are less likely to be released by appro- priate stimuli (for review see Grosvenor and Mena, 1992). Thus, prolactin must not only be synthesised but transformed before exocytotic release, giving consid- erable scope for differential effects. Transformation is inhibited by dopamine and promoted by dopamine an- tagonists including haloperidol and domperidone (Grosvenor et al., 1984). Dopamine may act directly on lactotrophs in the anterior pituitary by being released at terminals in the median eminence, but evidence from co-culture experiments suggests that cells in the intermediate lobe of the pituitary are additionally necessary for prolactin release (Ellekmann et al., 1991) and that the intermediate lobe may be tonically suppressed by dopamine. Furthermore, short portal vessels arising from the posterior pituitary are also believed to carry trophic factors controlling prolactin synthesis (Hyde et al., 1987) although their nature and origin are unclear at present.

Dopamine and Thyroid Hormone Release (See Fig. 3c)

Thyrotrophin releasing hormone (TRH) occurs in a wide variety of neurons, but those of most functional significance in controlling release of thyroid stimulating hormone (TSH) are in the parvocellular component of the paraventricular nucleus. They are stimulated to release TRH into the pituitary portal system by norad- renergic (and possibly adrenergic) neurons (Liposits et al., 1987; Shioda et al., 1986). It is believed that TRH release can be inhibited by dopaminergic neurons mak- ing presynaptic contact in the tubero-infimdibulum (Dieguez et al., 1984; Andersson and Eneroth, 1987).

It is, therefore, possible that bromocriptine might op- erate through changes in the pituitary-thyroid axis. Injections of thyroxine (T4) or triiodothyronine (T3) reduce the porphyrin content of female hamster Harde- rian glands and have a similar effect in castrated males, preventing the expected rise in porphyrin content (Hoffman et al., 1989,1990). The administration of antithyroid treatments such as methimazole and

KC104 have produced conflicting effects between different studies, but, reviewing these, Buzzell and Me- nendez-Pelaez (1992) concluded that “both hyperthy- roid and hypothyroid conditions tend to reduce porphyrin concentrations.”

Some studies have reported Harderian gland receptors for TSH, but the position for T3 and T4 receptors is unknown (see review by Buzzell & Menendez-Pelaez, 1992). Type II 5-deiodinase (5-D), an enzyme which is necessary for converting T4 to T3, is present in the Harderian glands of laboratory rats and hamsters (Del- gad0 et al., 1988; Guerrero et al., 1987,1989) but has not been identified in mice (Rubio et al., 1991). This enzyme is believed to be important in maintaining intracellular T3 levels in tissues which are thyroid hormone dependent, and activity is elevated in hypothy- roidism (Silva and Leonard, 1985). The enzyme has a diurnal rhythm believed to be controlled by adrenergic mechanisms, since the P-agonist isoprotenerol induces a rise which mimics that occurring in the dark phase. Vaughan and Guerrero (1992) have commented that this suggests a different control mechanism in the Harderian gland compared to the pineal; in the former, adrenergic neurons end chiefly on blood vessels, while in the latter they terminate directly on secretory cells.

Other Trophic Hormone Systems Dopamine has effects on the release of growth hor-

mone (Cronin et al., 1984; Lindstrom and Ohlsson, 19871, a-melanocyte stimulating hormone (Goudreau et al., 19921, and possibly corticotrophins (Gudelsky et al., 1989), but it is not clear a t present whether these can affect the Harderian gland. It is also possible that bromocriptine may be affecting some other mediator such as somatostatin which has been found in the Harderian gland (Aguilera and Catt, 1984; Puig-Dom- ingo et al., 1988). A direct effect ofbromocriptine on the gland must also be considered. For example, bromocriptine could reduce free calcium (Elsholtz et al., 1991) necessary for exocytosis or the availability of ace- tylcholine (Cummings, 1991) necessary for myoepithe- lial cell contraction. This could interfere with porphyrin secretion mechanisms. Alternatively, stimulation of D2 receptors by bromocriptine could alter CAMP levels and thus affect a variety of enzymatic and synthetic activities (Arunakaren et al., 1990; Elsholtz et al., 1991; Schettini et al., 1983; Swennen and Denef, 1982). Whether the Harderian gland possesses D1/D2 receptors is not known.

Given the complexity and multiplicity of the neuroendocrine systems mentioned above in which dopamine may play a (largely inhibitory) role, it is difficult with- out additional information to ascribe the effects of bromocriptine described by Buzzell et al. (1989) to any one system or trophic hormone. Severing the pituitary stalk or transplanting the pituitary to a peripheral site deprives the pituitary of its hypothalamic control, and its chief secretory product becomes prolactin which is normally inhibited by dopamine. Either of these proce- dures results in feminisation of the male hamster Harderian gland in that type I1 cells disappear and porphyrin is synthesised (Buzzell et al., 1992). How- ever, the obvious disruption of other neuroendocrine


T A B U 3. The frequency of tubules (Trtbs) containing type II cells, the frequency of type II cells, the fiquency of polytubular complexes (PTC) per unit area, and the frequency of mast cells per unit area in intact control male hamsters (Con), i n males castrated for 6 weeks (Cast), in males castrated for 6 weeks

and receiving the dopamine agonist bromncriptine (Cast + bro), in castrated males receiving both bromncriptine and proluctin (Cast + bro + pro). and in castmted males receiving prolactin alone

(Cast + pro)'

h e II cells Number Tubs (%) Cells (%F PTC Mast cells

Con 8 86.2 t 5.8 35.8 ? 2.6 0.96 2 0.22 0.35 ? 0.09 cast 8 LO* t 0.5 0.3* 2 0.2 O M * 2 0.13 3.27* 2 0.77 Cast + bro 6 100 62.2* t 2.5 1.07 2 0.11 1.10 ? 0.31 Cast + bro + pro 5 63.2 t 22.5 10.6* 2 4.5 0.02* 2 0.01 2.95* ? 0.68 Cast + pro 4 O* O* 0.01* 2 0.01 2.59* 2 0.60 F <0.001 <0.001 <0.001 <0.001

'For details of regimens, see text. All figures are means 5 sem. *Differs from control, P < 0.05.

TABLE 4. Levels of porphyrin (nmollg tissue) and the activity of the porphyrin rate-limiting enzyme G-aminolaeuulinate (ALA)-synthase (nmol ALA fonnedlhlg

protein) i n the Harderian glands of control female golden hamsters (Con), in ouariectomised females (Our), in ouariectomised females receiving the dopamine

agonist bromocriptine by continuous infmion (Our + bro), and in ovariectomised females receiving bromocriptine plus proluctin (Ovx + bro + pro)'

Porphyrin ALA-s Number eontent activity

Con 7 5,307 2 1,161 2,385 t 333 ovx 5 4,113 ? 466 1,185 2 56* Ovx + bro 5 2,867 t 658* 700 t 157* Ovx + bro + pro 5 6,869 2 1,106 1,614 t 267 F P < 0.05 P < 0.001

'For details of regimens, see text. All figures are means It sem. *Differs from control values, P < 0.05.

TABLE 5. The percentage of tubules containing type II cells (9% tubs), the percentage of type II cells (% cells), the frequency of polytubular complexes (PTC) per square micrometer, and the frequency of mast cells

per square millimeter in sections of the Harderian glands of control female golden humters (Con), i n ouariectomised females (Oux), in ovariectomised females receiving the dopamine agonist bromocriptine by continuous infusion (Oux + bro), and in ovariectomised females receiving bromocriptine plus prolactin by

continuous infusion (Our + bro + pro)'

TvDe II cells Number (% Tubs) (% Cells) €TC Mast cells

Con 7 0 0 0 10.16 ? 1.93 OVX 5 0 0 0 33.17* ? 2.99

4.66* ? 0.25 Ovx + bro 5 loo* Ovx + bro + pro 5 11.0* ? 1.64 4.0 t 0.5 0.01 ? 0.01 8.53 ? 0.32 F P < 0.001 P < 0.001 P < 0.001 P < 0.001

38.2* t 2.6 0.40* t 0.10

'For details of regimens, see text. All figures are means 2 sem. *Differs from control values, P i 0.05.

control mechanisms (particularly of gonadal and adre- nal activity) could have equally dramatic effects on the Harderian gland.

EFFECTS OF BROMOCRIPTINE AND PROLACTIN ON GLAND STRUCTURE AND

PORPHYRIN SYNTHESIS In recent experiments conducted on castrated male

and ovariectomised female hamsters over 6 week peri- ods, the continuous infusion of bromocriptine (1 mg/ day) by osmotic minipumps led to the suppression of porphyrin biosynthesis and the masculinisation of Harderian gland structure, changes which can be pre-

vented by the simultaneous implantation of minipumps containing prolactin (6 I.U. = 0.2 mg/day).

Experiments Using Intact and Castrated Male Hamsters

Porphyrin Biosynthesis Within the Hanlerian Glands (See Table 2). Intact (control) males possessed very little porphyrin within the Harderian gland and had low ALA-s activity. Conversely, males castrated for 6 weeks had considerably elevated synthesis. However, if castrated males received bromocriptine, they showed no such rise. The simultaneous administration of prolactin to the bromocriptine-treated


castrates resulted in significantly higher porphyrin levels and enzyme activity. Finally, castrated males treated with prolactin alone showed the highest rise in porphyrin levels, and animals so treated had porphyrin levels substantially higher than any other group.

Morphological Changes (See Table 3). Type I1 cells, typical of the male gland, are reduced after castration to negligible numbers. This decrease is not only prevented by bromocriptine, but castrates treated with this dopamine agonist possessed significantly more type I1 cells than normal males. The expected fall in type I1 cell numbers does occur if castrates are simul- taneously treated with prolactin. Similar changes also occur in polytubular complexes (which decrease in castrates and are preserved with bromocriptine treatment but decrease in castrated males given both bromocriptine and prolactin) and in mast cells (which increase in number in castrates and do not increase in bromocriptine-treated castrates but increase in bromocriptine- treated castrates which also receive prolactin).

Experiments Using Intact and Ovariectomised Female Hamsters

Porphyrin Bioeynthesis Within the Harderian Gland (See Table 4). Ovariectomy alone does not result in a marked decrease in porphyrin within the female Harderian gland, although ALA-s activity is significantly reduced. However, if ovariectomised females are given bromocriptine, the reduction in both enzyme activity and porphyrin content is marked. Both porphyrin content and enzyme activity are maintained in bromocriptine-treated ovariectomised females which also receive prolactin.

Morphological Changes (See Table 5). Neither intact nor ovariectomised females normally exhibit type I1 cells; nor do the gland epithelial cells contain polytubular complexes. However, if ovariectomised females receive bromocriptine, the frequency of type I1 cells is comparable to male glands (cf. Table 3). The simultaneous administration of prolactin reduces the frequency of type I1 cells and polytubular complexes towards female levels. Similarly, mast cells are significantly increased by ovariectomy (as previously reported by Spike et al., 1986a) but significantly decreased in ovariectomised females receiving bromocriptine; mast cell numbers are maintained at normal control levels in bromocriptine-treated ovariectomised females which also receive prolactin.

It thus appears that prolactin can act as an additional physiological control on gland structure and porphyrin biosynthesis in the hamster. Its effect on porphyrinogenic activity in other species is not known. However, in the rat, neither bromocriptine nor prolactin treatment had significant effects on lysosomal enzymes, while hypophysectomy alone altered acid phosphatase, a-mannosidase, and P-glucuronidase activities (Vaughan et al., 1988); these authors concluded that growth hormone (GH) might be a more important control for lysosomal enzymes.

It is clear from the present study that prolactin can increase porphyrin biosynthesis in castrated males when administered alone and will counteract the sup- pressive effects of bromocriptine. These effects can be

demonstrated on both porphyrin content and the activity of the rate-limiting enzyme ALA synthase. This suggests that the effects of bromocriptine reported by Buzzell et al. (1989) are probably manifested through prolactin inhibition and, if so, that prolactin may be as important a “coarse tuning” on gland structure and biosynthesis as androgens. It remains unclear why the gland should have two such major controlling mechanisms.

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Hormones and the control of porphyrin biosynthesis and structure in the hamster Harderian gland

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