CLIN. AND EXPER. HYPER.-THEORY AND PRACTICE, A 1 2 ( 4 ) , 631-645 (1990)

FURTHER STUDIES ON THE NORADRENERGIC AND THE R E N I N - ANGIOTENSIN SYSTEMS I N THE B R A I N OF THE RAT.

Maria L u i s a Kurn jek , Maria Rosa T r o l l i e t , L i d i a M i k u l i c , N i d i a Basso .

I n s t i t u t o d e I n v e s t i g a c i o n e s C a r d i o l 6 g i c a s . F a c u l t a d d e Medic ina . U n i v e r s i d a d d e Buenos A i r e s .

Marcel0 T . d e Alvea r 2 2 7 0 . 1 1 2 2 Buenos A i re s . A r g e n t i n a .

KEY WORDS: Norep inephr ine Ang io tens inogen Blood P r e s s u r e Catecholamines H e a r t Blood V e s s e l s P a r g y l i n e Cereb rosp i n a 1 F l u i d Monoamine o x i d a s e I n h i b i t o r s .

ABSTRACT

P r e v i o u s i n v e s t i g a t i o n s have shown t h a t d e p l e t i o n of b r a i n n o r e p i n e p h r i n e (NE) induced by chemica l sympa- thectomy r e s u l t e d i n s i g n i f i c a n t changes i n t h e c e n t r a l r e n i n - a n g i o t e n s i n sys t em. The pu rpose of t h e p r e s e n t work was t o i n c r e a s e t h e NE c o n c e n t r a t i o n i n t h e c e n t r a l ne rvous sys tem (CNS) i n o r d e r t o a n a l y z e i t s e f f e c t on t h e p e p t i d e r g i c complex and on t h e blood p r e s s u r e (BP) l e v e l s . T r e a t e d r a t s were g i v e n t h e f o l l o w i n g d r u g s i n t h e d r i n k i n g w a t e r : 1 -dopa (12 m g / r a t / d a y ) , c a r b i d o p a ( 6 m g / r a t / d a y ) and p a r g y l i n e (10 m g / r a t / d a y ) d u r i n g 2 5 d a y s . BP was d e t e r m i n e d , b lood and c e r e b r o s p i n a l f l u i d (CSF) samples were o b t a i n e d . The CNS was d i s s e c t e d i n t o s e v e r a l a r e a s . N E , ang io tens inogen . (AoC) and r e n i n con- c e n t r a t i o n (RC) were de t e rmined i n t h e b r a i n parenchyma; AoC was e v a l u a t e d i n CSF and plasma samples . Pharmaco-

631

Copyright 0 1990 by Marcel Dekker, Inc.

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l o g i c a l t r e a t m e n t r e s u l t e d i n a n h y p o t e n s i v e e f f e c t and , a t t h e same t ime, an i n c r e a s e o f NE i n t h e CNS (abou t 1 0 0 % ; p C O . 0 0 0 5 ) . These changes were accompanied by a s i g n i f i c a n t d e c r e a s e i n t h e p e r i p h e r a l and c e n t r a l AoC. These r e s u l t s add new e v i d e n c e t o t h e p o s t u l a t e d r e l a - t i o n s h i p between t h e s e two i m p o r t a n t r e g u l a t o r y sys tems invo lved i n c a r d i o v a s c u l a r c o n t r o l .

I N T R O D U C T I O N

A l l t h e components of t h e r e n i n - a n g i o t e n s i n sys tem have been i d e n t i f i e d i n t h e b r a i n of r a t s and dogs (1 - 7 ) and s i g n i f i c a n t i n f o r m a t i o n has been a f f o r d e d on t h e p h y s i o l o g i c a l r o l e and t h e r e l e v a n c e of t h e b r a i n enzy- matic sys tem. Recent s t u d i e s u s i n g m o l e c u l a r b i o l o g i c a l t e c h n i q u e s have shown t h a t r e n i n and a n g i o t e n s i n o g e n genes a r e e x p r e s s e d i n t h e b r a i n and i n o t h e r o rgans ( 8 - 1 1 ) . Even though t h e i n f l u e n c e of some f a c t o r s on t h e r e n i n and a n g i o t e n s i n o g e n gene e x p r e s s i o n h a s been demons t r a t ed i n some t i s s u e s ( 1 2 - 1 4 ) , r e s e a r c h i n t o t h e mechanisms r e l a t e d t o t h e r e g u l a t i o n and p h y s i o - p a t h o l o g i c a l meaning o f t h e c e n t r a l p e p t i d e r g i c system i s s t i l l i n i t s i n i t i a l s t a g e s .

I n p r e v i o u s e x p e r i m e n t s w e found t h a t b r a i n a n g i o - t e n s i n o g e n c o n c e n t r a t i o n (AoC) i n c r e a s e d i n a c o r r e s - ponding manner i n c e r e b r o s p i n a l f l u i d (CSF) (15) and i n c e r t a i n a r e a s of t h e c e n t r a l ne rvous sys t em (CNS) ( 6 ) . I n f u r t h e r s t u d i e s we obse rved t h a t chemica l sympathec- tomy w i t h i n t r a c e r e b r o v e n t r i c u l a r ( i . c . v . ) a d m i n i s t e r e d 6-hydroxydopamine (6-HODA), which caused a marked d e - p l e t i o n of b r a i n n o r e p i n e p h r i n e (NE), produced an i m - p o r t a n t increment i n a n g i o t e n s i n o g e n c o n c e n t r a t i o n i n CSF and i n some r e g i o n s of t h e CNS w i t h o u t i nduc ing changes i n t h e p e r i p h e r a l p e p t i d e . r g i c o r n o r a d r e n e r g i c sys tems ( 1 6 ) . Moreover , P r i n t z e t a l . ( 1 7 ) have found t h a t b r a i n a n g i o t e n s i n o g e n may be i n c r e a s e d by c e n t r a l n o r a d r e n e r g i c d e p l e t i o n . These r e s u l t s c o u l d s u g g e s t

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NORADRENERGIC AND RENIN SYSTEMS IN THE BRAIN 633

some k ind of c o r r e l a t i o n between t h e l e v e l s of AoC and t h o s e of NE c o n c e n t r a t i o n i n t h e b r a i n .

P r e s e n t expe r imen t s were des igned t o f u r t h e r e l u c i - d a t e t h e l i n k between t h e c e n t r a l p e p t i d e r g i c and n o r - a d r e n e r g f c sys tems. A pharmaco log ica l combina t ion of a

NE p r e c u r s o r , a p e r i p h e r a l i n h i b i t o r of deca rboxy lase and a monoamine ox idase (MAO) i n h i b i t o r were used t o i n c r e a s e t h e l e v e l s of t h e c e n t r a l NE c o n c e n t r a t i o n . In t h i s c o n d i t i o n , t h e c e n t r a l and p e r i p h e r a l n o r a d r e - n e r g i c and r e n i n - a n g i o t e n s i n sys tems were e v a l u a t e d t o i n v e s t i g a t e t h e p o s s i b l e r e l a t i o n s h i p between them and a l s o t h e i r p u t a t i v e r o l e i n blood p r e s s u r e (BP) r e g u l a t i o n .

MATERIALS AND METHODS

Male Wis t a r r a t s of t h e CHbbTHOM a l b i n o s t r a i n weighing about 250 g were used . The an ima l s were f e d a s t a n d a r d P u r i n a r a t chow ( C a r g i l l Argen t ina ) and were o f f e r e d t a p water t o d r i n k ; t h e y were kep t i n a n a u t o - m a t i c a l l y l i g h t e d room (from 0 7 . 0 0 A . M . t o 0 7 . 0 0 P.M.) a t a c o n s t a n t t empera tu re (22 - + l ° C ) . The r a t s were d i v i d e d i n t o two g roups : 1 . - Con t ro l a n i m a l s : t e n normal r a t s d r i n k i n g t a p water.

The amount of wa te r g i v e n d a i l y was e q u a l i z e d t o t h a t consumed by t h e expe r imen ta l g roup , d u r i n g 25 days .

2 . - T r e a t e d an ima l s : s i x normal r a t s d r i n k i n g t a p wa te r c o n t a i n i n g t h e f o l l o w i n g d r u g s : 1-dopa ( 1 2 m g / r a t / d a y ) , c a r b i d o p a ( 6 mg / ra t /day ) and p a r g y l i n e ( 1 0 mg/ ra t /day ) d u r i n g 25 days . Glucose ( 3 % ) was added t o t h e d r i n k i n g s o l u t i o n t o f a c i l i t a t e wa te r i n t a k e

Experimental an ima l s d rank less t h a n c o n t r o l r a t s , t h u s t h e l a t t e r r e c e i v e d o n l y a d a i l y amount of wa te r equa l t o t h a t consumed by t h e t r e a t e d r a t s t h e day b e f o r e .

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Blood p r e s s u r e w a s de t e rmined by t a i l p l e thysmogra - phy i n a l l t h e c o n s c i o u s animals b e f o r e beg inn ing t h e exper iment and 2 4 days l a t e r . A t t h e end o f t h e

e x p e r i m e n t a l p e r i o d , b lood samples were o b t a i n e d by c u t t i n g t h e t i p of t h e t a i l i n t h e c o n s c i o u s a n i m a l s ; plasma was s e p a r a t e d and k e p t f r o z e n f o r t h e e v a l u a t i o n of plasma r e n i n a c t i v i t y (PRA) and plasma a n g i o t e n s i n o - gen c o n c e n t r a t i o n . A f t e r w a r d s , t h e a n i m a l s were a n e s t h e - t i z e d w i t h sodium p e n t o b a r b i t a l ( 4 0 mg/kg, i . p . ) and CSF samples were o b t a i n e d from t h e c i s t e r n a magna as p r e v i o u s l y d e s c r i b e d (1 5 ) . The an ima l s were p r o c e s s e d immedia te ly t o remove t h e b r a i n (18) ; t h e f o l l o w i n g a r e a s were d i s s e c t e d : hypotha lamus , c e r e b r a l c o r t e x , c e r e b e l l u m , b r a i n stem, medu l l a o b l o n g a t a and s p i n a l c o r d . The t i s s u e s were l o n g i t u d i n a l l y c u t i n t o two s i m i l a r p o r t i o n s ; one h a l f was used f o r NE e v a l u a t i o n and t h e o t h e r f o r a n g i o t e n s i n o g e n and r e n i n - l i k e concen- t r a t i o n d e t e r m i n a t i o n s . The h e a r t , t h e m e s e n t e r i c a r t e - r ies and t h e a d r e n a l g l a n d s were a l s o e x c i s e d t o measure c a t e c h o l a m i n e s c o n c e n t r a t i o n .

-c Dete rmina t ion of Ang io tens inogen and R e n i n - l i k e Concen- t r a t i o n . --

The hypotha lamus , medu l l a o b l o n g a t a and s p i n a l co rd were homogenized i n 2 0 0 u l of 8 mM e t h y l e n e d i a m i n e t e t r a - a c e t i c a c i d (EDTA) i n s a l i n e ; 2 5 % homogenates were p repa red w i t h t h e o t h e r b r a i n a r e a s . A l l t h e homogenates were c e n t r i f u g e d a t 1 0 0 0 g f o r 3 0 m i n u t e s a t 4°C. The s u p e r n a t a n t s were s e p a r a t e d and an a l i q u o t was incuba ted i n t h e p r e s e n c e of a n op t ima l c o n c e n t r a t i o n of a n g i o t e n - s i n a s e i n h i b i t o r s (19) w i t h e i t h e r a n e x c e s s of semi- p u r i f i e d a n g i o t e n s i n o g e n from plasma of nephrec tomized r a t s , f o r r e n i n - l i k e c o n c e n t r a t i o n d e t e r m i n a t i o n o r a n e x c e s s of hog k idney r e n i n , f o r AoC e v a l u a t i o n . Measure-

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NORADRENERGIC AND RENIN SYSTEMS IN THE BRAIN 635

ment of plasma and CSF a n g i o t e n s i n o g e n c o n c e n t r a t i o n was per formed by i n c u b a t i o n of c o n v e n i e n t l y d i l u t e d

samples w i t h a n e x c e s s of hog k idney r e n i n ; PRA w a s de t e rmined by i n c u b a t i n g t h e samples w i t h a d e q u a t e a n g i o t e n s i n a s e i n h i b i t o r s ( 2 0 ) . I n c u b a t i o n s were con- d u c t e d i n t h e p r e s e n c e of 0 . 0 4 M t r i s c h l o r i d e b u f f e r (pH: 7 .2) f o r 60 m i n u t e s f o r b r a i n , plasma and CSF AoC and d u r i n g 3 h f o r r e n i n - l i k e c o n c e n t r a t i o n and PRA a t 37°C. The p r e s e n c e of a n g i o t e n s i n I (Ang I ) i n a l l samples w a s e v a l u a t e d by radioimmunoassay (RIA) (Pha- d e b a s , Pharmacia D i a g n o s t i c s , Uppsa la , Sweden). S i n c e a n g i o t e n s i n a s e i n h i b i t o r s i n t e r f e r e d w i t h t h e b i n d i n g r e a c t i o n o f t h e R I A , t h e same amount of these compounds added t o t h e samples were i n c o r p o r a t e d t o each Ang I s t a n d a r d c u r v e t o compensate f o r b i n d i n g a l t e r a t i o n s .

D e t e r m i n a t i o n of Cat e c holam i n e s

A f t e r d i s s e c t i o n , a l l t h e samples were f r o z e n i n a n a c i d i c s o l u t i o n (HC104-EDTA-NaZS03, 10 -1 -1 ) a t - 7 O O C .

The t i s s u e s were homogenized i n t h e same s o l u t i o n and c e n t r i f u g e d a t 1 0 0 0 g f o r 30 minu tes a t 4°C. The s u p e r - n a t a n t s were used t o s e p a r a t e t h e c a t e c h o l a m i n e s emplo- y i n g ch romatograph ic a lumina columns. The e l u a t e d amines were evaluated f l u o r i m e t r i c a l l y i n t h e a d r e n a l g l a n d s by t h e C r o u t ' s method (27) and NE c o n c e n t r a t i o n i n a l l o t h e r t i s sues was de t e rmined u s i n g t h e method d e s c r i b e d by L a v e r t y and T a y l o r ( 2 2 ) .

The d e t e c t i o n l i m i t f o r t h e R I A i s C 40 pg Ang I / m l , p r e c i s i o n between a s s a y s i s 5 . 5 % and w i t h i n a s s a y s i s 1 0 % . The s e n s i t i v i t y f o r NE i s < 20 ng/ml and t h e i n t e r and i n t r a - a s s a y v a r i a b i - l i t y i s 9 % and 7 . 2 % r e s - p e c t i v e l y . Data a r e e x p r e s s e d a s means + S.E.M. D i f f e - r e n c e s between g roups were e s t a b l i s h e d by t h e S t u d e n t ' s "t" t e s t .

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NORADRENERGIC AND RENIN SYSTEMS IN THE BRAIN 637

TABLE 2 .

c o n t r o l and t r e a t e d r a t s . Catecholamines c o n c e n t r a t i o n i n p e r i p h e r a l t i s s u e s o f

C o n t r o l T issue Sample Catecholamine

Heart NE (ng /g ) 432.3 - + 1 2 . 2

(1 0)

Mesenteric Artery NE (ng/g)3012.5 + 137.4 - (1 0)

Adrenal Gland NE (ng /g ) 168.6 - + 9.8 E (ng/g) 566.6 - + 38.2

(1 0) * p c o . 0 5 ; ** p c o . 0 2 ; *** pco.01.

T r e a t e d

*** 699.5 - + 29.5

(6)

3898.0 - + 378.8**

(6) ** * 123.2 - + 14.4

429.5 + 48.5 - ( 6 )

The body we igh t o f t r e a t e d r a t s was lower t h a n t h a t of c o n t r o l a n i m a l s . A t t h e end o f t h e s t u d y t r e a t e d r a t s weighed 245.5 - + 6 . 3 g and c o n t r o l r a t s 304 .9 - + 8 . 4 g , p C 0 . 0 0 1 . No o t h e r p h y s i c a l d i f f e r e n c e s were obse rved .

b e f o r e b e g i n n i n g t h e t r e a t m e n t (109.2 - + 2 . 7 v s 112 .2 - +

3 . 2 mmHg); a t t h e end of t h e expe r imen t t r e a t e d r a t s showed lower l e v e l s of BP t h a n any of t h e two i n i t i a l g r o u p s : 96.2 - + 5.5 mmHg and a l s o lower l e v e l s t h a n t h e c o n t r o l a n i m a l s : 113.7 - + 1 . 5 mmHg, pC 0 . 0 1 .

T r e a t e d r a t s p r e s e n t e d a s i g n i f i c a n t i n c r e a s e i n NE c o n c e n t r a t i o n i n a l l t h e a r e a s of t h e CNS. R e s u l t s a r e p r e s e n t e d i n Tab le 1 . A t t h e same time, t h e r e was a s i g n i f i c a n t d e c r e a s e i n AoC i n t h e CSF and i n t h e b r a i n parenchyma. Renin c o n c e n t r a t i o n was n o t a l t e r e d by t h e p r e s e n t pha rmaco log ica l t r e a t m e n t . R e s u l t s a r e a l s o i n c l u d e d i n T a b l e 1 .

S y s t o l i c BP was s i m i l a r i n b o t h g roups of r a t s

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NE c o n c e n t r a t i o n i n t h e m e s e n t e r i c a r t e r y and i n t h e h e a r t was s i g n i f i c a n t l y h i g h e r i n t r e a t e d r a t s w h i l e NE and e p i n e p h r i n e were d i m i n i s h e d i n t h e a d r e - n a l g l a n d s (Tab le 2) of e x p e r i m e n t a l a n i m a l s .

PRA l e v e l s were h i g h e r t h a n t h o s e u s u a l l y obse rved w i t h ou r methodology of b lood samples e x t r a c t i o n ; t h e y showed a tendency t o h i g h e r v a l u e s i n t r e a t e d r a t s (C: 24.6 - + 2 . 3 v s T r . : 35.1 - + 7 . 1 ng Ang I / m l / h , pCO.1) t h a t d i d n o t a t t a i n s t a t i s t i c a l s i g n i f i c a n c e ; AoC i n plasma was d i m i n i s h e d by t h e t r e a t m e n t (C: 5 5 0 . 0 - + 6 2 . 0 v s T r . : 307.7 - + 1 8 . 6 ng Ang I / m l , p e 0 . 0 2 ) .

DISCUSSION

Manipu la t ion of c e n t r a l n o r e p i n e p h r i n e l eve l s h a s been used as a t o o l t o a n a l y z e p o s s i b l e i n t e r a c t i o n s between t h e n o r a d r e n e r g i c and r e n i n - a n g i o t e n s i n sys t ems i n t h e c e n t r a l n e r v o u s sys t em and t h e i r r o l e on b lood p r e s s u r e r e g u l a t i o n i n t h e r a t . T h i s aim h a s been a c h i e - ved by t r e a t i n g t h e an ima l s w i t h : 1 . - a p e r i p h e r a l i n - h i b i t o r of a r o m a t i c aminoac id d e c a r b o x y l a s e ( c a r b i d o p a ) t h a t does n o t p e n e t r a t e t h e b r a i n ; 2 . - l a r g e d o s e s of 1-dopa , a p r e c u r s o r o f NE t h a t c r o s s e s t h e b lood b r a i n b a r r i e r and c o u l d e n t e r t h e c e n t r a l b i o s y n t h e t i c p a t h - ways of t h e a r o m a t i c amine w i t h o u t t h e i n t e r v e n t i o n of t h e r a t e l i m i t i n g enzyme, t y r o s i n e h y d r o x y l a s e and ; 3 . - a n i n h i b i t o r of monoamine o x i d a s e (MAO), t h e c y t o - p l a smic m e t a b o l i z i n g enzyme of monoamines, i n c l u d i n g NE ( p a r g y l i n e ) . Leve l s of c e n t r a l NE c o n c e n t r a t i o n were s i g n i f i c a n t l y e l e v a t e d a f t e r c h r o n i c t r e a t m e n t of t h e r a t s . Increments as h i g h a s 1 0 0 % were r e c o r d e d i n a l l b r a i n a r e a s and a l s o i n t h e s p i n a l c o r d . Higher NE l e v e l s were s p e c i a l l y p r e s e n t i n t h e h y p o t h a l a m u s , b r a i n stem and medul la o b l o n g a t a . Norad rene rg ic n u c l e i s p e c i - f i c a l l y invo lved i n BP r e g u l a t i o n and c a r d i o v a s c u l a r

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c o n t r o l a r e l o c a t e d i n t h e s e a r e a s . P r e s e n t r e s u l t s conf i rm a p r e v i o u s o b s e r v a t i o n on changes i n t h e b r a i n s tem NE c o n c e n t r a t i o n ( 2 3 ) .

Augmented NE c o n c e n t r a t i o n i n t h e b r a i n was accom- pan ied by a s i g n i f i c a n t d e c r e a s e i n b lood p r e s s u r e i n normal a n i m a l s . T h i s o b s e r v a t i o n was s imilar t o t h o s e r e p o r t e d by o t h e r a u t h o r s i n W i s t a r Kyoto r a t s r e c e i v i n g e i t h e r t h e same t r e a t m e n t (23) o r o n l y p a r g y l i n e (24 ) . An i n v e r s e r e l a t i o n s h i p between b r a i n s t e m NE l eve l s and blood p r e s s u r e was obse rved i n r a t s t r e a t e d w i t h MA0 i n h i b i t o r s ( 2 3 , 24) s u g g e s t i n g t h a t t h e accumula- t i o n of t h e amine i n t h e CNS was r e l a t e d t o t h e d r o p i n b lood p r e s s u r e . Moreover, NE i n j e c t i o n i n t o t h e n u c l e u s t r a c t u s s o l i t a r i i d e c r e a s e d b lood p r e s s u r e i n t h e r a t ( 2 5 ) . The BP l o w e r i n g e f f e c t o f t h e p r e s e n t t r e a t m e n t has been a s c r i b e d t o a c e n t r a l n o r a d r e n e r g i c modu la t ion o f b lood p r e s s u r e i n t h e r a t , r e l a t e d ma in ly t o t h e s t i m u l a t i o n of a l p h a 2 r e c e p t o r s l o c a t e d i n t h e b r a i n s tem a n d / o r medul la o b l o n g a t a . T h i s e f f e c t cou ld be s imi l a r t o t h a t o b t a i n e d w i t h c l o n i d i n e ( 2 6 ) o r a l p h a methyldopa ( 2 7 ) .

d e s i g n was t o a n a l y z e t h e a l t e r a t i o n s induced by h i g h c e n t r a l NE l e v e l s on t h e b r a i n r e n i n - a n g i o t e n s i n compo- n e n t s . P r e s e n t r e s u l t s have shown a g e n e r a l i z e d d e c r e a s e i n a n g i o t e n s i n o g e n c o n c e n t r a t i o n i n t h o s e a r e a s where NE was augmented. I n acco rdance w i t h p r e v i o u s r e s u l t s (16) d e p l e t i o n o f b r a i n NE by chemica l sympathectomy induced by i . c . v . 6-HODA r e s u l t e d i n a s i g n i f i c a n t i n c r e a s e i n AoC i n c e r e b r o s p i n a l f l u i d and i n b r a i n parenchyma. Fur the rmore , t h e r e is e v i d e n c e of a d e - c r e a s e d t u r n o v e r r a t e of NE i n t h e b r a i n stem of r a t s w i t h d o c a - s a l t h y p e r t e n s i o n ( 2 8 ) . I n t h e s e a n i m a l s an i n c r e a s e d r e n i n - a n g i o t e n s i n a c t i v i t y due t o h i g h e r l e - v e l s of a n g i o t e n s i n o g e n c o n c e n t r a t i o n i n t h e b r a i n has

The main i n i t i a l i n t e r e s t o f t h i s e x p e r i m e n t a l

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been proposed ( 6 ) . The p r e s e n t d a t a g i v e s u p p o r t t o t h e t h e s i s of an i n v e r s e r e l a t i o n s h i p between t h e n o r - a d r e n e r g i c and t h e p e p t i d e r g i c systems i n t h e b r a i n .

BP could be due t o d iminished p e r i p h e r a l sympathe t ic d r i v e induced by an i n c r e a s e d NE c o n c e n t r a t i o n a t a l p h a Z r e c e p t o r s i n t h e c e n t r a l nervous sys tem, and/ o r b l u n t i n g of a b r a i n a n g i o t e n s i n system which has been hypothes ized t o modulate n o r a d r e n e r g i c t o n e ( 2 9 ) and be r e l a t e d t o no rep inephr ine s t o r e s ( 1 7 ) .

p h e r a l components of bo th systems a r e a l s o of p a r t i - c u l a r i n t e r e s t . The p e r i p h e r a l r e n i n - a n g i o t e n s i n s y s - tem may have been s t i m u l a t e d ( a s r e f l e c t e d i n a h ighe r PRA w i t h a lowered ang io tens inogen c o n c e n t r a t i o n ) due t o t h e d iminished wa te r i n t a k e i n bo th groups . Simul- t a n e o u s l y , NE was i n c r e a s e d a t t h e ne rve t e r m i n a l s of t h e h e a r t and blood v e s s e l s of t r e a t e d an imals . As f a r a s w e know, t h e e f f e c t of t h e p r e s e n t combinat ion of drugs on t h e p e r i p h e r a l NE c o n c e n t r a t i o n has n o t been p r e v i o u s l y ana lyzed . O the r s have r e p o r t e d t h a t MA0 i n h i b i t i o n r e s u l t e d i n an i n c r e a s e d NE c o n c e n t r a t i o n i n t h e r a t ( 3 0 ) and gu inea p i g (31) h e a r t and i n blood v e s s e l s of bo th s p e c i e s (31 , 3 2 ) . F u r t h e r , c h r o n i c t r ea tmen t w i t h p a r g y l i n e and l -dopa r e s u l t e d i n i n - c r eased NE i n c a t c o e l i a c g a n g l i a ( 3 3 ) . However, MA0 i n h i b i t o r s have a l s o been r e p o r t e d t o i n h i b i t NE syn- t h e s i s i n t h e h e a r t and blood v e s s e l s (31) p o s s i b l y by feedback r e g u l a t i o n of e l e v a t e d NE. Our r e s u l t s would, t h e r e f o r e , sugges t dec reased r e l e a s e of NE r e s u l t i n g i n i n c r e a s e d s t o r e s . There i s no c lear ex- p l a n a t i o n f o r t h e observed d e c r e a s e i n a d r e n a l medu- l l a r y ca techolamines s t o r e s . Dairman e t a l . , ( 3 4 ) r e p o r t e d reduced a d r e n a l t y r o s i n e hydroxylase a c t i - v i t y a f t e r l -dopa t r e a t m e n t . Th i s may be due e i t h e r

I n t h e p r e s e n t s t u d y , t h e observed lowering of

The r e s u l t s ob ta ined i n t h e a n a l y s i s of t h e p e r i -

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t o f e e d b a c k i n h i b i t i o n ( f rom s y n t h e t i z e d NE o r ep i - n e p h r i n e ) o r , p o t e n t i a l l y , t o d i s p l a c e m e n t of NE f rom s t o r a g e s i t e s .

s e n t p h a r m a c o l o g i c a l t rea tment induced a s i g n i f i c a n t NE accumula t ion i n t h e c e n t r a l ne rvous sys t em, h e a r t and b lood v e s s e l s w h i l e i t produced a d e - c r e a s e i n BP t h a t c o u l d be due t o d i m i n i s h e d p e r i - p h e r a l s y m p a t h e t i c d r i v e . I n c r e a s e d NE c o n c e n t r a - t i o n i n t h e b r a i n i s a s s o c i a t e d w i t h s i g n i f i c a n t d e p l e t i o n of c e n t r a l a n g i o t e n s i n o g e n c o n c e n t r a t i o n . T h i s a l t e r a t i o n was o p p o s i t e t o t h a t obse rved a f t e r NE d e p l e t i o n by i . c . v . a d m i n i s t r a t i o n of 6-HODA ( 1 6 ) . Our r e s u l t s s u p p o r t t h e e x i s t e n c e of a l i n k between b r a i n n o r a d r e n e r g i c and s e l e c t e d p e p t i d e r g i c s y s t e m s .

I n summary, it c a n be conc luded t h a t t h e p r e -

ACKNOWLEDGEMENTS --

The p r e s e n t i n v e s t i g a t i o n was accompl ished w i t h Gran t 3057200/85 from t h e Conse jo Nac iona l d e I n v e s t i - g a c i o n e s C i e n t i f i c a s y T6cn icas (CONICET) d e A r g e n t i n a . We g r a t e f u l l y acknowledge t h e s k i l l f u l t e c h n i c a l a s s i s - t a n c e of M s Graciela D o t t a .

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1 9 7 1 ; 1 7 7 : 1 1 9 - 1 2 6 . -

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3 3 . - P u i g , M . , Wakadem, A . R . , K i r p e k a r , S.M.- E f f e c t on t h e s y m p a t h e t i c ne rvous system of c h r o n i c t r e a t m e n t w i t h p a r g y l i n e and 1-DOPA.. J . P h a r m a c o 1 . E ~ ~ . T h e r . 1 9 7 2 ; 1 8 2 : 1 3 0 - 1 3 4 . -

3 4 . - Dairman, W . , U d e n f r i e n d , S . - Dec rease i n a d r e n a l t y r o s i n e h y d r o x y l a s e and i n c r e a s e i n n o r e p i n e p h r i - ne s y n t h e s i s i n r a t s g i v e n 1 -dopa . S c i e n c e . 1 9 7 1 ; 1 7 1 : 1 0 2 2 - 1 0 2 4 . -

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