European Journal of Pharmacology, 182 (1990) 199- 202
199
Elsevier EJP 20650 Short communication
G A B A B receptor-mediated contractile effects resistant to tetrodotoxin in isolated cat ileum Nevena Pencheva, Kalina Venkova and Radomir
Radomirov
Institute of Physiology, Bulgarian Academy of Sciences, A cad. G. Bonchev Str. bl. 23, 1113 Sofia, Bulgaria
Received 11 April 1990, accepted 1 May 1990
The effects of 7-aminobutyric acid (GABA) and GABAergic drugs were studied on longitudinal strips from cat terminal ileum prepared after removing the myenteric plexus. GABA and baclofen exerted concentration-dependent contractile effects. Muscimol was ineffective, and bicuculline did not antagonize the effect of GABA. The complete elimination of the neural input to the smooth muscle cells by tetrodotoxin failed to prevent the action of GABA and baclofen. Pharmacological analyses of the effects indicated the existence of GABA B receptors on the smooth muscle cells in the longitudinal layer of cat terminal ileum. Ileum (cat isolated terminal); GABA (y-aminobutyric acid); Smooth muscle effect; Baclofen; G A B A a receptors
1. Introduction
Recently, 3,-aminobutyric acid ( G A B A ) has b e e n r e c o g n i z e d as an a u t o n o m i c n e u r o t r a n s m i t t e r in the enteric n e r v o u s system where G A B A A a n d G A B A B r e c e p t o r s are f o u n d (Jessen et al., 1987). In general, bicuculline-sensitive G A B A A receptors, which are c o u p l e d to a p i c r o t o x i n i n - s e n s i t i v e C1- i o n o p h o r e , a n d the less u n d e r s t o o d bicuculline-insensitive G A B A B receptors, which are indep e n d e n t of C I - , m e d i a t e the c o n t r a c t i l e a n d rel a x a n t effects of G A B A on intestinal activity. E l e c t r o p h y s i o l o g i c a l a n d p h a r m a c o l o g i c a l experim e n t s have revealed that G A B A r e c e p t o r s are l o c a t e d on n e u r o n a l e l e m e n t s (Jessen et al., 1987; F a r g e a s et al., 1988). These r e c e p t o r s are t h o u g h t n o t to be p r e s e n t on the gut m u s c u l a t u r e , since G A B A has n o direct effects on the c o n t r a c t i l i t y of gut s m o o t h muscle. However, the presence of
Correspondence to: R. Radornirov, Institute of Physiology, Bulgarian Academy of Sciences, Acad. G. Bonchev Str. bl. 23, 1113 Sofia, Bulgaria.
G A B A r e c e p t o r s on secretory cells a n d on s m o o t h muscle cells of the o v i d u c t a n d uterus has also been d o c u m e n t e d (Erdo, 1985). In o u r study, we used d e n e r v a t e d intestinal p r e p a r a t i o n s to c h a r a c t e r i z e p h a r m a c o l o g i c a l l y the effects of G A B A on s p o n t a n e o u s m e c h a n i c a l activity.
2. Materials and methods 2.1. S m o o t h muscle preparations
M a l e a d u l t cats weighing 3.5-4.0 kg were used in the experiments. T h e a n i m a l s were killed after they h a d been a n a e s t h e t i s e d with c h l o r a l o s e (100 m g / k g ) . A 7-8 c m length of the t e r m i n a l ileum (3 c m a b o v e the ileocaecal sphincter) was r e m o v e d a n d flushed with K r e b s s o l u t i o n o f the following c o m p o s i t i o n ( m M ) : NaC1 120, KC1 5.9, N a H C O 3 15.4, N a H 2 P O 4 1.2, MgC12 1.2, CaC12 2.5, glucose 11.5. Segments of the t e r m i n a l ileum ( a p p r o x i m a t e l y 3 c m long) were cut a l o n g the m e s e n t e r i c insertion a n d stretched in a large Petri dish con-
0014-2999/90/$03.50 © 1990 Elsevier Science Publishers B.V. (Biomedical Division)
200 taining Krebs solution. After the mucosa and submucosa had been removed, the circular muscle layer with Auerbach's plexus was gently separated under a low power binocular microscope. Plexusfree longitudinal strips about 3 m m wide and 18-20 m m long were selected and cut from the ileum sections containing the plexus with fine scissors. Some of the strips were not completely free of nerves because plexus-retaining areas, although damaged, were detected histochemically. The strips were mounted at an initial tension of 0.5 g in a 10 ml organ bath containing Krebs solution at 3 6 ° C bubbled with 5% CO 2 and 95% 02 . Before the experiments started, the preparations were allowed to equilibrate for 50 min during which the solution was changed several times. Mechanical activity of the strips was recorded by means of a strain gauge (M 1000 B, Microtechna) connected to a recording device (Type-175, Kutest).
2.2. Pharmacological analysis and statistics Mechanical responses to G A B A and to a G A B A B agonist were determined as a function of concentration both in untreated strips and on the background produced by antagonists. Drugs remained in contact with the tissue for not more than 3 min and were then washed out. Desensitization was examined by measuring responses to drugs without intervening washings. Intervals of 20-30 min (with washings) between single doses of G A B A and baclofen were sufficient to prevent desensitization. Responses are expressed as percentage of the m a x i m u m response elicited by the agonist. The antagonists were used at one concentration only. Concentration-effect curves were analysed with linear regression and the test for parallelism described by Tallarida and Murray (1981). Student's t-test at P < 0.05 was used to calculate the 95% confidence limits for the ECs0 values and the slopes of the regression lines.
2.3. Drugs All drug solutions were prepared before the experiment by dissolving the agents in distilled
water, except bicuculline, which was dissolved in 0.1 N HC1 and adjusted to p H 6.5 with 0.1 N N a O H . The drug volumes varied with the experiments, but they were never more than 1% of the bath volume. The following drugs were used: G A B A (Merck), ( + ) - b a c l o f e n (Research Biochemicals Inc.), muscimol (Fluka AG), bicuculline (Fluka AG), tetrodotoxin (Sankyo).
3. Results
Spontaneous mechanical activity was observed in longitudinal strips from cat terminal ileum after the equilibration period. Under our experimental conditions, motility was characterized by a steady state tone and rhythmic phasic contractions. G A B A (5-1000 /~M) and the G A B A B receptor agonist, baclofen (5-500 /~M), exerted concentration-dependent contractile effects. The response had two components - a pronounced increase in phasic contraction amplitudes and an elevation in basal tone (fig. 1A). The stimulation induced by G A B A or baclofen was sustained, achieving a maximum within 2 min, and decreased only gradually to the control level. In order to avoid desensitization, drugs were washed out after the m a x i m u m effect had been reached. Usually the two components of the responses developed at the same time, but there were cases when the tonic contraction preceded the increase in spontaneous phasic activity. When G A B A (500/~M) was left in the organ bath for 5 min, during which the resting tension recovered, the repeated application of G A B A (1000 /~M) or baclofen (500 /xM) was ineffective. Thus specific and cross-desensitization occurred between G A B A and baclofen. Desensitization and cross-desensitization were reversible after washing within 25-30 min. The G A B A A receptor agonist, muscimol, at concentrations up to 500 # M was ineffective, or produced only a weak enhancement of the phasic component without changing the tone. Concentration-effect curves for G A B A and baclofen were obtained in untreated strips and in the presence of tetrodotoxin. Concentration-effect curves for G A B A were also studied after treatment with bicuculline. The ECs0 value for baclo-
201 BACLOFEN 500 pM
GABA
I
Sml~
h
4. Discussion
BIcUCULLINE
TTX
I0 ~M
Fig. 1. Longitudinal plexus-free strips from cat terminal ileum. Effects of GABA and baclofen on the spontaneous mechanical activity in untreated strips (A), and in the presence of bicuculline (B) and tetrodotoxin (C).
fen determined in untreated strips was more than two times lower that for G A B A (table 1). The selective blocker of G A B A A receptors, bicuculTABLE 1 The contractile effects of GABA and baclofen in control and pharmacologically pretreated strips. TTX, tetrodotoxin, c.l., confidence limits. Correl. coeff., correlation coefficient of regression. n GABA
GABA after bicuculline (50 ~M) GABA after TTX (1 ~M) Baclofen Baclofen after TTX (1/~M)
ECs0 (/aM) (95% c.1.)
Slope (95% c.l.)
Correl. coeff.
14
26.16 (23.82-30.57)
40.53 (33.53-47.53)
0.97
12
27.53 (24.35-35.83)
37.97 (25.97-49.97)
0.95
9
33.10 (33.88-40.73) 11.71 (10.22-12.50) 11.11
35.71 (30.51-40.73) 32.86 (21.52-44.20) 40.70
0.99
(9.41-12.22)
(39.80-51.60)
15 9
line, at a dose of 50 /~M did not antagonize the effects of G A B A or baclofen (fig. 1B), and had no significant effect on the ECs0 values for G A B A (table 1). Tetrodotoxin failed to prevent the GABA- and baclofen-induced stimulatory effects (fig. 1C). The ECs0 values were similar when estimated in untreated strips and in the presence of tetrodotoxin (table 1). After applying the test for parallelism, no significant differences were found between the slopes of the five concentration-effect curves.
0.87 0.96
High endogenous G A B A levels have been demonstrated in the enteric nervous system of various mammals (Jessen et al., 1986; 1987; Davanger et al., 1987; Erdo, 1985). It has been found that GABAergic neurons project not only within the myenteric plexus but also into the circular and longitudinal coat of the gut wall (Furness et al., 1989). The role of G A B A in the transmission of smooth muscle is not clear, but it is considered possible that the effects of G A B A released from terminals within the muscle require the presence or co-release of another transmitter. The present experiments show that, in plexusfree longitudinal strips from cat terminal ileum, G A B A produces pronounced contractile effects. Baclofen mimics the stimulatory effect of GABA, including the tonic and phasic components and its sustained nature. However, when the ECs0 values of baclofen and G A B A in untreated strips were compared, baclofen was found to be more potent. On the other hand, the ECs0 values for G A B A in the absence and presence of bicuculline did not differ significantly indicating that the contraction produced by G A B A may be mediated through G A B A a receptors. Furthermore, cross-desensitization was observed between G A B A and baclofen, suggesting a common site of action. The lack of action of muscimol, as well as the bicuculline insensitivity of the G A B A - and baclofen-induced effects, excludes GABAA-mediated activity in strips with absent or damaged myenteric plexus. On the contrary, in longitudinal strips from the cat terminal ileum retaining the myenteric
202 plexus, GABA A receptors mediate neurogenic c h a n g e s of t h e s p o n t a n e o u s a c t i v i t y ( u n p u b l i s h e d d a t a ) , as t h e y d o in the c a t c o l o n ( T a n i y a m a et al., 1987). The resistance of GABA- and baclofen-induced r e s p o n s e s to t e t r o d o t o x i n , w h i c h e l i m i n a t e s p a r t i c i p a t i o n o f n e u r o n a l t r a n s m i t t e r release m e c h a n i s m s ( K a t z a n d M i l e d i , 1968), s u g g e s t s t h a t the observed GABA B effects are not mediated through a u t o n o m i c nerves. T h u s the c o n t r a c t i l e effects in the l o n g i t u d i n a l l a y e r c o u l d b e d u e to a n o n - n e u ronal excitatory, postsynaptic mechanism. Taking i n t o a c c o u n t the l a c k o f s i g n i f i c a n t d i f f e r e n c e s b e t w e e n t h e s l o p e s of the c o n c e n t r a t i o n - e f f e c t curves, it m a y b e a s s u m e d t h a t o n e a n d the s a m e t y p e o f p o s t s y n a p t i c G A B A r e c e p t o r s is i n v o l v e d in the c o n t r a c t i l e e f f e c t s o f G A B A a n d b a c l o f e n in t h e cat i l e u m . O u r d a t a p r o v i d e e v i d e n c e t h a t in the l o n g i t u d i n a l l a y e r o f t h e cat t e r m i n a l i l e u m , as in t h e r a b b i t F a l l o p i a n t u b e ( E r d o et al., 1984), G A B A exerts a direct contractile action through GABA a receptors o n s m o o t h m u s c l e cells.
References Davanger, S., O.P. Ottersen and J. Storm-Mathisen, 1987, Immunocytochemical localization of GABA in cat myenteric plexus, Neurosci. Lett. 73, 27.
Erdo, S.L., M. Riesz, E. Karpati and L. Szporny, 1984, GABA B receptor-mediated stimulation of the contractility of isolated rabbit oviduct, European J. Pharmacol. 99, 33. Erdo, S.L., 1985, Peripheral GABAergic mechanisms, Trends Pharmacol. Sci. 5, 205. Fargeas, M.J. Fioramonti and L. Bueno, 1988, Central and peripheral action of GABA A and GABA B agonists on small intestine motility in rats, European J. Pharmacol. 150, 163. Furness, J.B., D.C. Trussell, S. Pompolo, J.C. Bornstein, B.E. Maley and J. Storm-Mathisen, 1989, Shapes and projections of neurons with immunoreactivity for gammaaminobutyric acid in the guinea-pig small intestine, Cell Tissue Res. 256, 293. Jessen, K.R., J.M. Hills and M.J. Saffrey, 1986, lmmunohistochemical demonstration of GABAergic neurons in the enteric nervous system, J. Neurosci. 6, 1628. Jessen, K.R., R. Mirsky and J.M. Hills, 1987, GABA as an autonomic neurotransmitter: studies on intrinsic GABAergic neurons in the myenteric plexus of the gut, Trends Neurosci. 10, 255. Katz, B. and R. Miledi, 1968, The effect of local blockade of motor nerve terminals, J. Physiol. 199, 729. Tallarida, R.J. and R.B. Murray, 1981, In: Manual of Pharmacological Calculation with Computer Programs, eds. R.J. Tallarida and R.B. Murray (Springer-Verlag, New York Inc.). Taniyama, K., N. Saito, Y. Miki and C. Tanaka, 1987, Enteric gamma-aminobutyric acid-containing neurons and the relevance to motility of the cat colon, Gastroenterology 93, 519.
199
Elsevier EJP 20650 Short communication
G A B A B receptor-mediated contractile effects resistant to tetrodotoxin in isolated cat ileum Nevena Pencheva, Kalina Venkova and Radomir
Radomirov
Institute of Physiology, Bulgarian Academy of Sciences, A cad. G. Bonchev Str. bl. 23, 1113 Sofia, Bulgaria
Received 11 April 1990, accepted 1 May 1990
The effects of 7-aminobutyric acid (GABA) and GABAergic drugs were studied on longitudinal strips from cat terminal ileum prepared after removing the myenteric plexus. GABA and baclofen exerted concentration-dependent contractile effects. Muscimol was ineffective, and bicuculline did not antagonize the effect of GABA. The complete elimination of the neural input to the smooth muscle cells by tetrodotoxin failed to prevent the action of GABA and baclofen. Pharmacological analyses of the effects indicated the existence of GABA B receptors on the smooth muscle cells in the longitudinal layer of cat terminal ileum. Ileum (cat isolated terminal); GABA (y-aminobutyric acid); Smooth muscle effect; Baclofen; G A B A a receptors
1. Introduction
Recently, 3,-aminobutyric acid ( G A B A ) has b e e n r e c o g n i z e d as an a u t o n o m i c n e u r o t r a n s m i t t e r in the enteric n e r v o u s system where G A B A A a n d G A B A B r e c e p t o r s are f o u n d (Jessen et al., 1987). In general, bicuculline-sensitive G A B A A receptors, which are c o u p l e d to a p i c r o t o x i n i n - s e n s i t i v e C1- i o n o p h o r e , a n d the less u n d e r s t o o d bicuculline-insensitive G A B A B receptors, which are indep e n d e n t of C I - , m e d i a t e the c o n t r a c t i l e a n d rel a x a n t effects of G A B A on intestinal activity. E l e c t r o p h y s i o l o g i c a l a n d p h a r m a c o l o g i c a l experim e n t s have revealed that G A B A r e c e p t o r s are l o c a t e d on n e u r o n a l e l e m e n t s (Jessen et al., 1987; F a r g e a s et al., 1988). These r e c e p t o r s are t h o u g h t n o t to be p r e s e n t on the gut m u s c u l a t u r e , since G A B A has n o direct effects on the c o n t r a c t i l i t y of gut s m o o t h muscle. However, the presence of
Correspondence to: R. Radornirov, Institute of Physiology, Bulgarian Academy of Sciences, Acad. G. Bonchev Str. bl. 23, 1113 Sofia, Bulgaria.
G A B A r e c e p t o r s on secretory cells a n d on s m o o t h muscle cells of the o v i d u c t a n d uterus has also been d o c u m e n t e d (Erdo, 1985). In o u r study, we used d e n e r v a t e d intestinal p r e p a r a t i o n s to c h a r a c t e r i z e p h a r m a c o l o g i c a l l y the effects of G A B A on s p o n t a n e o u s m e c h a n i c a l activity.
2. Materials and methods 2.1. S m o o t h muscle preparations
M a l e a d u l t cats weighing 3.5-4.0 kg were used in the experiments. T h e a n i m a l s were killed after they h a d been a n a e s t h e t i s e d with c h l o r a l o s e (100 m g / k g ) . A 7-8 c m length of the t e r m i n a l ileum (3 c m a b o v e the ileocaecal sphincter) was r e m o v e d a n d flushed with K r e b s s o l u t i o n o f the following c o m p o s i t i o n ( m M ) : NaC1 120, KC1 5.9, N a H C O 3 15.4, N a H 2 P O 4 1.2, MgC12 1.2, CaC12 2.5, glucose 11.5. Segments of the t e r m i n a l ileum ( a p p r o x i m a t e l y 3 c m long) were cut a l o n g the m e s e n t e r i c insertion a n d stretched in a large Petri dish con-
0014-2999/90/$03.50 © 1990 Elsevier Science Publishers B.V. (Biomedical Division)
200 taining Krebs solution. After the mucosa and submucosa had been removed, the circular muscle layer with Auerbach's plexus was gently separated under a low power binocular microscope. Plexusfree longitudinal strips about 3 m m wide and 18-20 m m long were selected and cut from the ileum sections containing the plexus with fine scissors. Some of the strips were not completely free of nerves because plexus-retaining areas, although damaged, were detected histochemically. The strips were mounted at an initial tension of 0.5 g in a 10 ml organ bath containing Krebs solution at 3 6 ° C bubbled with 5% CO 2 and 95% 02 . Before the experiments started, the preparations were allowed to equilibrate for 50 min during which the solution was changed several times. Mechanical activity of the strips was recorded by means of a strain gauge (M 1000 B, Microtechna) connected to a recording device (Type-175, Kutest).
2.2. Pharmacological analysis and statistics Mechanical responses to G A B A and to a G A B A B agonist were determined as a function of concentration both in untreated strips and on the background produced by antagonists. Drugs remained in contact with the tissue for not more than 3 min and were then washed out. Desensitization was examined by measuring responses to drugs without intervening washings. Intervals of 20-30 min (with washings) between single doses of G A B A and baclofen were sufficient to prevent desensitization. Responses are expressed as percentage of the m a x i m u m response elicited by the agonist. The antagonists were used at one concentration only. Concentration-effect curves were analysed with linear regression and the test for parallelism described by Tallarida and Murray (1981). Student's t-test at P < 0.05 was used to calculate the 95% confidence limits for the ECs0 values and the slopes of the regression lines.
2.3. Drugs All drug solutions were prepared before the experiment by dissolving the agents in distilled
water, except bicuculline, which was dissolved in 0.1 N HC1 and adjusted to p H 6.5 with 0.1 N N a O H . The drug volumes varied with the experiments, but they were never more than 1% of the bath volume. The following drugs were used: G A B A (Merck), ( + ) - b a c l o f e n (Research Biochemicals Inc.), muscimol (Fluka AG), bicuculline (Fluka AG), tetrodotoxin (Sankyo).
3. Results
Spontaneous mechanical activity was observed in longitudinal strips from cat terminal ileum after the equilibration period. Under our experimental conditions, motility was characterized by a steady state tone and rhythmic phasic contractions. G A B A (5-1000 /~M) and the G A B A B receptor agonist, baclofen (5-500 /~M), exerted concentration-dependent contractile effects. The response had two components - a pronounced increase in phasic contraction amplitudes and an elevation in basal tone (fig. 1A). The stimulation induced by G A B A or baclofen was sustained, achieving a maximum within 2 min, and decreased only gradually to the control level. In order to avoid desensitization, drugs were washed out after the m a x i m u m effect had been reached. Usually the two components of the responses developed at the same time, but there were cases when the tonic contraction preceded the increase in spontaneous phasic activity. When G A B A (500/~M) was left in the organ bath for 5 min, during which the resting tension recovered, the repeated application of G A B A (1000 /~M) or baclofen (500 /xM) was ineffective. Thus specific and cross-desensitization occurred between G A B A and baclofen. Desensitization and cross-desensitization were reversible after washing within 25-30 min. The G A B A A receptor agonist, muscimol, at concentrations up to 500 # M was ineffective, or produced only a weak enhancement of the phasic component without changing the tone. Concentration-effect curves for G A B A and baclofen were obtained in untreated strips and in the presence of tetrodotoxin. Concentration-effect curves for G A B A were also studied after treatment with bicuculline. The ECs0 value for baclo-
201 BACLOFEN 500 pM
GABA
I
Sml~
h
4. Discussion
BIcUCULLINE
TTX
I0 ~M
Fig. 1. Longitudinal plexus-free strips from cat terminal ileum. Effects of GABA and baclofen on the spontaneous mechanical activity in untreated strips (A), and in the presence of bicuculline (B) and tetrodotoxin (C).
fen determined in untreated strips was more than two times lower that for G A B A (table 1). The selective blocker of G A B A A receptors, bicuculTABLE 1 The contractile effects of GABA and baclofen in control and pharmacologically pretreated strips. TTX, tetrodotoxin, c.l., confidence limits. Correl. coeff., correlation coefficient of regression. n GABA
GABA after bicuculline (50 ~M) GABA after TTX (1 ~M) Baclofen Baclofen after TTX (1/~M)
ECs0 (/aM) (95% c.1.)
Slope (95% c.l.)
Correl. coeff.
14
26.16 (23.82-30.57)
40.53 (33.53-47.53)
0.97
12
27.53 (24.35-35.83)
37.97 (25.97-49.97)
0.95
9
33.10 (33.88-40.73) 11.71 (10.22-12.50) 11.11
35.71 (30.51-40.73) 32.86 (21.52-44.20) 40.70
0.99
(9.41-12.22)
(39.80-51.60)
15 9
line, at a dose of 50 /~M did not antagonize the effects of G A B A or baclofen (fig. 1B), and had no significant effect on the ECs0 values for G A B A (table 1). Tetrodotoxin failed to prevent the GABA- and baclofen-induced stimulatory effects (fig. 1C). The ECs0 values were similar when estimated in untreated strips and in the presence of tetrodotoxin (table 1). After applying the test for parallelism, no significant differences were found between the slopes of the five concentration-effect curves.
0.87 0.96
High endogenous G A B A levels have been demonstrated in the enteric nervous system of various mammals (Jessen et al., 1986; 1987; Davanger et al., 1987; Erdo, 1985). It has been found that GABAergic neurons project not only within the myenteric plexus but also into the circular and longitudinal coat of the gut wall (Furness et al., 1989). The role of G A B A in the transmission of smooth muscle is not clear, but it is considered possible that the effects of G A B A released from terminals within the muscle require the presence or co-release of another transmitter. The present experiments show that, in plexusfree longitudinal strips from cat terminal ileum, G A B A produces pronounced contractile effects. Baclofen mimics the stimulatory effect of GABA, including the tonic and phasic components and its sustained nature. However, when the ECs0 values of baclofen and G A B A in untreated strips were compared, baclofen was found to be more potent. On the other hand, the ECs0 values for G A B A in the absence and presence of bicuculline did not differ significantly indicating that the contraction produced by G A B A may be mediated through G A B A a receptors. Furthermore, cross-desensitization was observed between G A B A and baclofen, suggesting a common site of action. The lack of action of muscimol, as well as the bicuculline insensitivity of the G A B A - and baclofen-induced effects, excludes GABAA-mediated activity in strips with absent or damaged myenteric plexus. On the contrary, in longitudinal strips from the cat terminal ileum retaining the myenteric
202 plexus, GABA A receptors mediate neurogenic c h a n g e s of t h e s p o n t a n e o u s a c t i v i t y ( u n p u b l i s h e d d a t a ) , as t h e y d o in the c a t c o l o n ( T a n i y a m a et al., 1987). The resistance of GABA- and baclofen-induced r e s p o n s e s to t e t r o d o t o x i n , w h i c h e l i m i n a t e s p a r t i c i p a t i o n o f n e u r o n a l t r a n s m i t t e r release m e c h a n i s m s ( K a t z a n d M i l e d i , 1968), s u g g e s t s t h a t the observed GABA B effects are not mediated through a u t o n o m i c nerves. T h u s the c o n t r a c t i l e effects in the l o n g i t u d i n a l l a y e r c o u l d b e d u e to a n o n - n e u ronal excitatory, postsynaptic mechanism. Taking i n t o a c c o u n t the l a c k o f s i g n i f i c a n t d i f f e r e n c e s b e t w e e n t h e s l o p e s of the c o n c e n t r a t i o n - e f f e c t curves, it m a y b e a s s u m e d t h a t o n e a n d the s a m e t y p e o f p o s t s y n a p t i c G A B A r e c e p t o r s is i n v o l v e d in the c o n t r a c t i l e e f f e c t s o f G A B A a n d b a c l o f e n in t h e cat i l e u m . O u r d a t a p r o v i d e e v i d e n c e t h a t in the l o n g i t u d i n a l l a y e r o f t h e cat t e r m i n a l i l e u m , as in t h e r a b b i t F a l l o p i a n t u b e ( E r d o et al., 1984), G A B A exerts a direct contractile action through GABA a receptors o n s m o o t h m u s c l e cells.
References Davanger, S., O.P. Ottersen and J. Storm-Mathisen, 1987, Immunocytochemical localization of GABA in cat myenteric plexus, Neurosci. Lett. 73, 27.
Erdo, S.L., M. Riesz, E. Karpati and L. Szporny, 1984, GABA B receptor-mediated stimulation of the contractility of isolated rabbit oviduct, European J. Pharmacol. 99, 33. Erdo, S.L., 1985, Peripheral GABAergic mechanisms, Trends Pharmacol. Sci. 5, 205. Fargeas, M.J. Fioramonti and L. Bueno, 1988, Central and peripheral action of GABA A and GABA B agonists on small intestine motility in rats, European J. Pharmacol. 150, 163. Furness, J.B., D.C. Trussell, S. Pompolo, J.C. Bornstein, B.E. Maley and J. Storm-Mathisen, 1989, Shapes and projections of neurons with immunoreactivity for gammaaminobutyric acid in the guinea-pig small intestine, Cell Tissue Res. 256, 293. Jessen, K.R., J.M. Hills and M.J. Saffrey, 1986, lmmunohistochemical demonstration of GABAergic neurons in the enteric nervous system, J. Neurosci. 6, 1628. Jessen, K.R., R. Mirsky and J.M. Hills, 1987, GABA as an autonomic neurotransmitter: studies on intrinsic GABAergic neurons in the myenteric plexus of the gut, Trends Neurosci. 10, 255. Katz, B. and R. Miledi, 1968, The effect of local blockade of motor nerve terminals, J. Physiol. 199, 729. Tallarida, R.J. and R.B. Murray, 1981, In: Manual of Pharmacological Calculation with Computer Programs, eds. R.J. Tallarida and R.B. Murray (Springer-Verlag, New York Inc.). Taniyama, K., N. Saito, Y. Miki and C. Tanaka, 1987, Enteric gamma-aminobutyric acid-containing neurons and the relevance to motility of the cat colon, Gastroenterology 93, 519.
