Effect of some vasoactive peptides on the lower esophageal sphincter

Pharmacolo#ical Research Communications, Vol. 12, No. 10, 1980

965

EFFECT OF SOME VASOACTIVE PEPTIDES ON THE LOWER ESOPHAGEAL SPHINC-TER o

G. Coruzzi and G. Bertaccini I n s t i t u t e of Pharmacology, University of Parma, Parma, I t a l y

SUMMARY .Some vasoactive peptides ( a n g i o t e n s i n , .vasopressin,:.bradykinin,

substance P.) and neurotensin which, under many- respects,

behaves as a bradykinin, were tested for t h e i r a c t i v i t y on .the isolated

lower esophageal sphincter (LES) of rats and guinea

~.igs and were compared with acetylcholine.L..In the r a t preparation a l l

the above compounds showeda striking-spasmogenic, a c t i -

v i t y and appeared to be more potent than acetylcholine in terms -

.

. .

.

.

of threshold dose.but less effective...in terms Of. maximum .response. I n

the guinea ~pig, preparation a l l .the. peptides but vaso-

pressin and substance P, were endowed with relaxant properties and threshold doses were higher than in the r a t . was a common feature

Tachyphylaxis

in both species: however.the

lack of the

common i n h i b i t o r s to modify the effects of the peptides suggests a d i r e c t a c t i v i t y on the sphincteric muscle.lt is l i k e l y that some of the examined peptides may have a role in-the complex regulation of LES tone at least by i n t e r a c t i n g ' w i t h other humoral and / or nervous mediators. °

Supported by a grant from the CNR, Roma.

0031-6989/80/100965-09/~;02.00/0

© 1980 The Italian PharmacologicalSociety

Pharmacological Research Communications, VoL 12, No. 10, 1980

966

INTRODUCTION Vasoactive peptides are. known to affect the c o n t r a c t i l i t Y not only.of vascular, but also of extravascu'lar smooth muscle (Sander and Huggins,.-1972; Bertaccini, 1976; Bolton, 1979). Gastrointestifial muscle is very sensitive to these peptides which usually cause stimulation and. increase both tone and amplitude..of spontaneous movements. The effect may be d i f f e r e n t in the various areas of the alimentary canal (Regoli et ai..,1974; Bertaccini, 1980): for this reason we decided .to investigate the action of some vasoactive peptides on-the: lower esophageal sphincter which is a d i f f e r e n t i a t e d ...

.

.

•

.

.....

area quite d i s t i n C t f r o m b o t h t h e esophageal body and the proximal- stomach (Christensen,l:975; LiPshutz and Cohen, 1977). Moreover, whereas the action of the gastrointestinal peptides on the LES has been thoroughly investigated ( f o r review, see .

.

. . .

Fisher and Cohen, 1980), that. of vasoactive peptides received •

_

(

so far l i t t l e

attention and ~nly preliminary reports are now

available .(Bertaccini et ai..,1980). Together with some "class i c a l " vasoactive peptides al"so neurotensin was considered since in several test preparations this peptide was found to behave as a bradykinin (Carraway and Leeman, 1973). The above peptides were studied on isolated LES preparations from rats and guinea pigs which-were selected for the easy a v a i l a b i l i t y of homogeneous animals. MATERIALSAND METHODS Male w i s t a r rats (weighing approximately 200 g)and guinea pigs :of:both sexes(weighing from 300. to 450 g ) w e r e k i l • .

.

- ~

,

~

~

. . . .

.

led bya blow on~:the neck;: segments 0.2 cm in length were tai kent.from:the lowest esophagus. They were opened by a longitu-

Pharmacological Research Communications, VoL 12, No. 10, 1980 a~nal

~ncislon

and used as a c i r c u l a r

967

muscle p r e p a r a t i o n •

the LES. S i l k

threads

were t i e d ' t o

both'ends

which were suspended, in a I0 ml bath fluid

and bubbl'ed w i t h

was f i x e d transducer nutrient 113;

connected

solution

KCl,

4~7;

2.5 and g l u c o s e After

215;

period

nistered.

cin

between

( I 0 mg/kg s u b c u t a n e o u s l y ) (according

Drugs used were:

NaHCO3,

was 1 . 5 g. each compound it

two subsequent a d m i n i s t r a t i o n s

angiotensin

LaboratoriesInc.

was • t r e a t e d w i t h

3 hr b e f o r e

to W h i t t l e ,

press'in , Sandoz) , s u b s t a n c e (Peninsula

(mM): NaCl,

in the Krebs s o l u t i o n . a n d

A sma1•l number o f r a t s

the e x p e r i m e n t

The-

2 - 4 min acco~di,ng to the~ dose admi-

Timeintervai

Was I0 min.

tension

isometric

1.2;

o f about 60 m i n ,

was added to the bath d i s s o l v e d to a c t f o r

MgSO4,

.

thestrip

Comerio).

composition

KH2P04, 1 . 2 ;

I I . • 5 ~ The i n i ' t i a l

an e q u i l i b r a t i o n

was a l l o w e d

(Basile,

had the f o l l o w i n g

CaCI2,

to=an

",,;

nutrient

One end o f

end Was attach'ed

to a recorder

.

the s t r lps

filled-with

oxygen at 37°C.

and the o t h e r

of

of

indometha-

the b e g i n n i n g

of

1978).

(C~"a). , vasopress:in

(kysB,vaso -

P, brady k i n i n and n e u r o t e n s i n California),

acetylcholine,

atro-

I

pine,

c h l o r p h e n i r a m i n e ; ~/ h e n t o l a m i n e

(Chiesi),

methysergide :(Sandoz):

(Fluka),

indomethacin

tetrodotoxin

(Sankyo).

preparations

from r a t s

RESULTS Results guinea

obtained

in

isolated

p i g s were d i f f e r e n t

qualitative

point

ved from T a b l e sensitive contracted

o f view

1 that. rat

than those

(Table

I).

In f a c t

preparations

from the guinea

by the p e p t i d e s :

was c o n t r a c t e d o n l y

from both a q u a n t i t a t i v e

and a be o b s e r -

were d e c i d e d l y

more

p i g and were u s u a l l y

conversely

by v a s o p r e s s i n

itcan

and

the guinea p i g LES

and substance

P but re-

+ + +

Vaso~ressin

Substance P

Neurotensin

: contraction;

- +

Bradykin~n

+

+

= relaxation;

Effect

*

*

*

*

*

= occurrence

5 . 9 x 10 -9

0 . 7 x 10 -7

1 0 x I0 -I0

Zi8 x I0- 8

2.9 x I0 -9

5 . 9 x 10 .8

2~2 x 10 -6

1 . 0 x 10 -6

2 . 8 x 10 -7

9 . 7 x 10-7

of tachyphylaxis-(T).

-

+

+

-

-

Inresno~a dose(t~)

Effect

Threshold dose~M) T

GUINEA PIG

RAT

*

*

*

*

T

ACTION OF SOME VASOACTIVE PEPTIDES AND NEUROTENSIN ON ISOLATED EES PREPARATIONS.

Angiotensin

Compound

Table I.

,9

Q~

c~ o

O0

CO

Pharmacolo.qical Research~Communications, Vol. 12, No. 10, 1980

•

B

Ach I

A1

969

B 0.3

0.5gl

N 0.3

lltlj o

/ I Achl Fig. 1

•

•

•

A3

B3

N3

Isolated LES p r e p a r a t i o n s ~ f ' . t h e r a t : ( o n ithe::itop) and ofl.ithe guinea p i g : . ( o n : t h e ~ t t o m ) , i A c h i ~ acetyicholi'ne; A.= angi6tensin; B = bradykinin; N = neurotensin, Doses are inlHg/ml Time ~ in imin On:the ordinata:tension •

"

"

i

*

'

"

i

.~

'

of thel trBnsducer in .grams, laxed by angiotensin, bradykinin and n~urotensin (Fig. I ) The high s e n s i t i v i t y of the rat LES allowed us to construct a complete dose-response curv..e, ~or a l l

the peptides examined,

u

as shown in Fig. 2 in which, f o r comparison,

also the dose-

response curve to acetylcholine is represented. I t is evident from Fig. 2 t h a t , with the exception of substance P, a l l

the

other peptides were more "potent" than a c e t y l c h o l i n e - i n terms of threshold stimulant doses. Conversely they a l l were less active than acetylcholine in terms of " e f f i c a c y " ( i . e . . m a x i -

970

Pharmacological Research Communications, VoL 12, No. 10, I980 Ac:h /

1D.

~

V

!

SP / / 1I

/

0.5.

/ O 0.3

Fig. 2

3

ng

30

3oo

I

1o

~g

1oo

Dose-response turves of d i f f e r e n t compounds on the isolated tower'.esophageal sphincter o f t h e

r a t . On the

ordinata, tension of the transducer in grams. Doses are in ng/ml or in ~g/ml. Each value refers to the mean of the values obtained from 5 to 10 experiments.

mum response). Unlike a c e t y l c h o l i n e , almost a l ] of the peptides were characterized by a marked tachyphylaxis which appeared at the second or t h i r d subsequent administration of the same peptide. Therefore the dose-response curves had to be performed by giving d i f f e r e n t doses in d i f f e r e n t preparations. Atropine, in doses (0.1 pg/ml) which completely prevented the maximum response to a c e t y l c h o l i n e , and tetrodotoxin (0.1 ~g/ml) added to the bath 15 min p r i o r to administration of the peptides ] e f t t h e i r effects absolutely unchanged. The same was

~armaco/ogicalResea~hCommun~ation~,~L'l~No.~l~

1980

971

true fol.lowing administration.~of~methisergide,~.phentolamine, chlorpheniramine or p r e t r e a t m e n t o f t h e animals, with~..indometha.cin DISCUSSZON Vasoactive peptides and neurot.en.sin affected LES..:moti!

lity

in isolated preparations of rats,and."guinea pigs and

t h e i r behaviour was d i f f e r e n t in the two animal species: the lower esophageal sphincter of the: rat was remarkably-more sensitive to the peptides than that of the guinea p i g ; - i n addition i,.t produced a contraction in response to a l l

the

peptides except bradykinin which exhibited a biphasic .response ( r e l a x a t i o n followed by c o n t r a c t i o n ) .

In the guinea pig re\ laxation was the most common feature, even in the case of an,.~.

.

~

.

giotensin, a peptide Which is known to contract a l l

the other

/

smooth muscle isolated preparations (Regoli e t a1.,1974). These observations t o g e t h e ~ t h e bradykinin (biphasic e f f e c t )

d i f f e r e n t behaviour of

and neurote~sin (only contraction)

on "the r a t LES, emphasize the importance of" the p a r a l l e l bioassay on both animal species in order to discriminate between peptides of the same family. The i n a b i l i t y of tetrodotoxin to modify the e f f e c t s of our peptides on the esophageal sphincter suggests an action which is not nervous mediated.

On the other hand the lack of

interf~ence of atropine, phentolamine, chlorpheniramine and methisergide and pretreatment with indomethacin

a]lowed us

to exclude an e f f e c t mediated by s t i m u l a t i o n of c h o l i n e r g i c , adrenergic, histaminergic or serotoninergic receptors or release of prostaglandins. Of course the possible release of

972

Pharmaco(ogicef Research.Communications. Vol. .12. No,..1O. 1980

other:unknown:mediators; which could explain the appearance o f tachyphylaxis, cannot be excluded on-the basis of..our experiments. Some g a s t r o i n t e s t i n a l hormones were found (Takayanagy and Kasuya, ~1977) to, contract..the LES o f the rat and guinea pig; however this~e.f:fect was scarcely evident in terms o f b o t h potency a n d e f f i c a c y . . This is apparently consistent with the idea that some o f , t h e peptides.examined in t h i s . i n v e s t i g a t i o n ( l i k e substance P and. neurotensin),

which were found to be pre-

sent.in-.the g a s t r o i n t e s t i n a l t r a c t and. are so:active even in very small amounts; have a role in .the physiological c o h t r o l of LES function i n . r a t s and guinea p i g s . . I t seems also plausible that-endogenous angiotensin and vasopressin may too a f f e c t LES m o t i l i t y at least in pathological conditions characterized by abnormal blood levels of these.peptides. Of course, in addi.tion to the e f f e c t of the single peptides, the possible interactions between them and/or other-humoral and nervous mediators should also be considered.

REFERENCES BERTACCINI, G. (1976). Pharmacol, Rev,, 28, 127'177. ,

BERTACCINI, G. (1980), In: G a s t r o i n t e s t i n a l Hormones (Jerzy Glass, G B., Ed.), pp. 315-341, Raven Press, New York. BERTACCINI, G , CORUZZI, G. and SCARPIGNATO, C. (1980). I n t e r . Symp. Medical and. Surg$cal .Problems of the Esophagus. Roma, May 7-9, BOLTON, T.B. (1979). Physiol, Rev., 59, 606-718.

Pharmacolo#ical Research Communications, VoL. 12,..No. I0, 1980

CARRAWAY, .R. Iand.LEEMAN, S..E. (197.3).~ J. B i o l . C;hem:,248, 6854-6861. CHRI.STENSEN., J- (7975).-Ann.. Rev.":Pharmacol.. ~, :15,:243--258. FISHER,.R.;S..-and COHEN.,-S.. (]980). in": GastrointestinalHormones (Jerzy~Glass, G.B., Ed.), Raven.:,Press, NewYork (in...press). LIPSHUTZ,".W. and COHEN,.S.~.(1971). Gastroenterology,: 61, 16-24. REGOLI, D., PARK, W.K. and RIOUX, .F. (.1974)../Pharmacol.; Rev., 25, 69-123. SANDER, G.E. and HbGGINS, C.G.- (1972). Ann/Rev. Pharma,co.l,, 1:2, 227-264,

TAKAYANAGIi. i. I and KASUYA, Y~, (1977),~ ~J.-Pharm. Pharmacol.~, '29', 559,560. WHITTLE, BIJI,R.

(1978). B r ,

.d. Pharmacol., 64. 438P.

973