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The action of morphine and naloxone on acid secretion by the rat isolated stomach
European Journal o f Pharmacology, 71 (1981) 135--138
135
© Elsevier/North-Holland Biomedical Press Short communication T H E A C T I O N O F M O R P H I N E A N D N A L O X O N E O N A C I D S E C R E T I O N BY T H E R A T ISOLATED STOMACH s. PAUL CANFIELD 1 and JAN SPENCER 2 1 Department o f Physiology, St. Mary's Hospital Medical School, London W2 1PG, and 2 Department o f Zoology, Westfield College, London NW3 7ST, U.K.
Received 14 November 1980, accepted 19 February 1981
S.P. CANFIELD and J.E. SPENCER, The action o f morphine and naloxone on acid secretion by the rat isolated stomach, European J. Pharmacol. 71 (1981)135--138. The action of morphine and naloxone on acid secretion by the rat isolated stomach has been studied. Morphine (10 -7 to 10 -4 M) had no effect upon spontaneous acid secretion. Morphine (10 -5 M) did not modify the acid output in response to sub-maximal stimulation by pentagastrin, histamine, bethanechol or isoprenaline. Naloxone (10 -5 M) was without effect on the response to pentagastrin or histamine. Our results suggest that opiate receptors do not modify acid secretion in this preparation. Morphine
Rat
Naloxone
Isolated
1. I n t r o d u c t i o n T h e r e have r e c e n t l y b e e n a n u m b e r o f p u b lications c o n c e r n e d w i t h t h e a c t i o n o f o p i a t e agonists o n gastric acid s e c r e t i o n in vivo. K o n t u r e k et al. ( 1 9 8 0 ) r e p o r t e d t h a t b o t h m o r p h i n e a n d M e t - e n k e p h a l i n caused dose d e p e n d e n t increases in acid s e c r e t i o n in b o t h H e i d e n h a i n p o u c h and fistula c o n s c i o u s d o g p r e p a r a t i o n s . T h e s e r e p o n s e s were i n h i b i t e d b y n a l o x o n e a n d also b y a t r o p i n e a n d m e t i a m i d e G a s c o i g n e et al. ( 1 9 8 0 ) f o u n d n o e f f e c t o f M e t - e n k e p h a l i n o n e i t h e r basal or stimulated acid s e c r e t i o n in fistula cats even at d o s e s high e n o u g h to cause a t o x i c r e a c t i o n . E1 Munshid et al. ( 1 9 8 0 ) r e p o r t e d t h a t Mete n k e p h a l i n i n h i b i t e d h i s t a m i n e , b u t n o t pentagastrin, s t i m u l a t e d acid s e c r e t i o n in conscious fistula rats. In h u m a n subjects, F e l d m a n et al. ( 1 9 8 0 ) f o u n d t h a t n a l o x o n e i n h i b i t e d acid s e c r e t i o n f o l l o w i n g a meal. H o w e v e r , m o r p h i n e also caused a r e d u c t i o n o f acid output. T h e s e studies d o n o t p r e s e n t a simple pict u r e o f the e f f e c t o f o p i a t e agonists o n acid
Acid
Stomach
secretion. In p a r t t h e d i f f e r e n c e s in t h e results m a y be d u e s i m p l y to species d i f f e r e n c e b u t m a y also be d u e t o t h e wide s p e c t r u m o f activity o f this class o f drugs in t h e b o d y . Opia t e agonists h a v e b e e n r e p o r t e d to increase gastric m u c o s a l b l o o d flow, cause the release o f h i s t a m i n e a n d gastrin and i n h i b i t the release o f a c e t y l c h o l i n e . T h u s o p i a t e s m a y m o d i f y acid s e c r e t i o n b y a v a r i e t y o f i n d i r e c t m e a n s r a t h e r t h a n b y a p r i m a r y a c t i o n on t h e parietal cell. We have t h e r e f o r e s t u d i e d t h e a c t i o n o f m o r p h i n e a n d n a l o x o n e o n acid s e c r e t i o n using an in vitro s t o m a c h p r e p a r a t i o n f r o m t h e r a t w h e r e s o m e o f t h e s e difficulties are absent.
2. Materials a n d m e t h o d s 2.1. P r e p a r a t i o n
S t o m a c h s were o b t a i n e d f r o m i m m a t u r e Wistar rats weighing 3 0 - - 5 0 g w h i c h h a d b e e n k e p t w i t h a lactating f e m a l e . A n i m a l s w e r e killed b y cervical d i s l o c a t i o n and t h e s t o m a c h
136
S.P. CANFIELD, J. SPENCER
rapidly r e m o v e d , cut along the lesser curvature, everted and washed with cold 0.9% saline solution. The s t o m a c h s were tied over the end o f a p e r s p e x t u b e (cross sectional area 1.1 cm 2) with the m u c o s a facing into the l u m e n o f t h e tube. The m u c o s a was b a t h e d by 5 ml o f u n b u f f e r e d saline gassed with 100% 02 and the serosal surface with 30 ml o f HCO~ b u f f e r e d saline (pH 7.4) gassed with 95% 02-5% CO2. T h e solutions were maintained at 34°C and their c o m p o s i t i o n was identical with t h a t previously used for the guinea pig s t o m a c h b y H o l t o n and S p e n c e r (1976). T h e mucosal saline was replaced at 15 min intervals, its pH m e a s u r e d and it was t i t r a t e d t o pH 7 with 20 mM NaOH.
over a 4 h period. All drugs were a d d e d to the serosal saline and left in c o n t a c t with the tissue for 1 h.
2.3. Drugs All drug dilutions were freshly m a d e on the day o f the e x p e r i m e n t and the following were used: pentagastrin (Peptavlon, ICI Ltd, U.K.), isoprenaline sulphate, histamine acid sulphate (Macarthys L t d , U.K.), b e t h a n e c h o l chloride ( G l e n w o o d L a b o r a t o r i e s Ltd, U.K.), Naloxo n e h y d r o c h l o r i d e (Narcan, W i n t h r o p e , U.K.) and m o r p h i n e sulphate (Evans Medical Ltd, U.K.).
2.4. Statistics 2.2. Expression of results A variable p r o p o r t i o n o f the i m m a t u r e rat s t o m a c h p r e p a r a t i o n consists of n o n - s e c r e t o r y r u m e n and part o f this was included within the area o f the perspex t u b e in some preparations w h e r e t h e r u m e n was relatively large. This r e d u c e d the area of s e c r e t o r y m u c o s a to less t h a n 1.1 cm 2. As it was n o t possible t o estimate a c c u r a t e l y the size o f this r e d u c t i o n t h e responses t o drug a d d i t i o n have been expressed as the s e c r e t o r y ratio (R) w h e r e R=
Average plateau response to drug Average spontaneous secretion
T h e average s p o n t a n e o u s secretion was obt a i n e d f r o m t h e rates for the t w o periods imm e d i a t e l y prior t o drug addition. Thus a value o f R = 1 indicates no stimulation whilst R = 2 shows a d o u b l i n g o f t h e rate o f secretion in t h e presence o f the drug. Thus, the spontaneous secretion (usually 2--4 ~ m o l H ÷ cm -2" h -1) has b e e n utilised as an i n d e x o f the a m o u n t o f s e c r e t o r y mucosa. S o m e absolute m e a n values o f acid secretion (response-spont a n e o u s in p m o l H ÷ cm -2 • h -1) with standard errors have b e e n included in the t e x t to facilit a t e c o m p a r i s o n o f this p r e p a r a t i o n with o t h e r published w o r k on isolated stomachs. C o n t r o l e x p e r i m e n t s s h o w e d t h a t the spont a n e o u s secretion did n o t change significantly
Comparisons b e t w e e n test s t o m a c h s and c o n t r o l s t o m a c h s were m a d e on the s e c r e t o r y ratios using the Mann-Witney U-test as this has the advantage t h a t it does n o t require assumptions a b o u t the statistical distribution o f the data.
3. Results
3.1. Action of morphine alone M o r p h i n e was a d d e d t o t h e s t o m a c h s b y c u m u l a t i v e increases in c o n c e n t r a t i o n over the range 10 -7 to 10 -4 M. T h e m e a n values o f R with standard errors are s h o w n in fig. 1. Each c o n c e n t r a t i o n was left in c o n t a c t for 1 h, t h e n washed o u t and i m m e d i a t e l y replaced by the higher c o n c e n t r a t i o n . N o n e o f the c o n c e n t r a tions used caused any significant change in the s p o n t a n e o u s secretion. T w o p o i n t concent r a t i o n - r e s p o n s e curves f o r histamine and isoprenaline have b e e n included for c o m p a r i s o n . In absolute values the response at 1 0 - 4 M m o r p h i n e was --0.17 + 0.11 p m o l H ÷ c m -2 • h -~, n o t significantly d i f f e r e n t f r o m zero. The responses to the higher c o n c e n t r a t i o n s o f hist a m i n e and isoprenaline were respectively 1.60 _+ 0.29; 3.19 _+ 0.48 p m o l H* cm -2 • h -1.
MORPHINE AND ACID OUTPUT BY RAT ISOLATED STOMACH
2.2 ~13 2.0 1.6 L2
137
TABLE 1 The effect of morphine (10 -6 M) and naloxone (10 -6 M) on secretagogue induced acid secretion in the rat isolated stomach. Test stomachs were preincubated with either morphine or naloxone for 1 h prior to addition of either pentagastrin (P), histamine (H), bethanechol (B) or isoprenaline (I) for a further hour. Control stomachs received only the secretagogues. Mean values with standard error and number of observations in brackets are shown. Probability values were obtained from Mann-Witney U-tests.
1.8
14
,.o ~_________~÷
Secretagogue
°'2,
,o
Drug
Concentration
,00
Secretory test
(microMolar)
Fig. 1. Secretory ratio R as ordinate plotted against log drug concentration as abscissa. Points are mean values with standard error bars. The number of observations (n) was 8 for all morphine points (A A). Two point curves for isoprenaline (e ¢,) and histamine (u --) are included for comparison and n is shown beside each point.
3.2. A c t i o n o f m o r p h i n e on o t h e r secretagogues We also i n v e s t i g a t e d t h e e f f e c t o f preincubating the stomachs with morphine ( 1 0 -6 M) f o r 1 h o n t h e s e c r e t o r y r e s p o n s e t o approximately half-maximal concentrations of either pentagastrin, histamine, bethanechol, or isoprenaline. Results were c o m p a r e d with identically treated control stomachs w h i c h d i d n o t h a v e m o r p h i n e . T h e r e s u l t s are s h o w n i n t a b l e 1A. C l e a r l y m o r p h i n e d i d n o t alter the response compared with the controls. For b e t h a n e c h o l the c o n t r o l secretion w a s 1 . 9 9 _+ 0 . 5 1 ~zmol H ÷ c m - 2 • h -1 a n d i n t h e presence of morphine 2.71 ± 0.86 pmol H ÷ c m - 2 . h-1.
3.3. A c t i o n o f n a l o x o n e Konturek e t al. ( 1 9 8 0 ) r e p o r t e d t h a t naloxone inhibited sub-maximal responses to p e n t a g a s t r i n a n d h i s t a m i n e as well as i n h i b i t ing o p i a t e - s t i m u l a t e d secretion in their cons c i o u s d o g s t u d i e s . We t h e r e f o r e e x a m i n e d the effect of naloxone (10-6M) on the
Ratio (R) control
Probability
0.24
A. Morphine (10 -6 M) P (10 -7 M)
1.65 (6) + 0.06
1.60 (6) + 0.09
H(4.4 × 1 0 - s M )
1.59 (8) + 0.04
1.47 (9) + 0.05
B (8 × 10 -6 M)
2.22 (6) + 0.31
2.02 (6) +- 0.39
0.35
I (6 × 10 -7 M)
1.79 (6) -+ 0.10
1.75 (6) + 0.12
0.16
1.48 (6) + 0.11
0.53
1.41 (6)
1.40 (6)
0.41
± 0.03
± 0.06
>0.1
B. Naloxone (10 -6 M) P (10 -7 M) H (4.4 × 10 -s M)
1.53 (5) + 0.10
response of the rat stomach to half-maximal concentrations of histamine and pentagastrin u s i n g t h e s a m e t e s t - c o n t r o l d e s i g n as a b o v e . T h e r e s u l t s are s h o w n i n t a b l e l B . N a l o x o n e had n o effect o n the responses at this c o n c e n tration. The absolute values for pentagastrin w e r e 1 . 9 0 + 0 . 2 7 p m o l H ÷ c m - : • h -1 f o r c o n t r o l s a n d 1 . 5 5 + 0 . 3 9 p m o l H * c m -2 . h -1 with naloxone. The spontaneous secretion for the p e n t a g a s t r i n c o n t r o l s was 3 . 1 6 + 0.48 p m o l H ÷ c m -2 • h -1 w h i l s t i n t h e p r e s e n c e o f n a l o x o n e it was 3 . 6 5 +_ 0 . 9 6 ~ m o l H + c m -2 • h -1. T h e y w e r e n o t s i g n i f i c a n t l y d i f f e r e n t .
4. D i s c u s s i o n Morphine did not have any effect on either s p o n t a n e o u s acid secretion or t h a t s t i m u l a t e d
138
by any of the four secretagogues used and naloxone was also without action on the two stimulants tested. In presenting negative results there is always the problem that they may arise from the use of poor preparations rather than indicating the behaviour of the system being studied. The stomachs used in this study secreted acid spontaneously and responded to four different stimulants. Although the number of observations was not large we have previously shown inhibitory effects in this preparation using other drugs in similar sized samples (Canfield et al., 1980). The use of secretory ratios rather than absolute values for the statistical tests would tend to favour positive results as the within treatment variance of the ratios tends to be lower than that with absolute values where no allowance is made for the variable a m o u n t of secretory mucosa. The secretory ratio might conceal changes in response if the spontaneous secretion were altered by the drug treatment under test in such a way that the ratio derived from this was unaltered. However, neither morphine nor naloxone altered spontaneous acid output. We therefore, feel confident that our negative results with morphine and naloxone indicate that there are no opiate receptors in the
S.P. CANFIELD, J. SPENCER
rat stomachs capable of modifying acid secretion. There remains the possibility that opiate receptors outside the stomach may influence acid secretion indirectly.
References Canfield, S.P., C.A. Price and J.E. Spencer, 1980, Noradrenaline and gastric acid secretion by the rat isolated stomach, Br. J. Pharmacol. 70,178P. El Munshid, H.A., R. Hakanson, G. Liedberg and F. Sundler, 1980, Effects of various gastrointestinal peptides on parietal cells and endocrine cells in the oxyntic mucosa of rat stomach, J. Physiol. 305,249. Feldman, M., J.H. Walsh and L.L. Taylor, 1980, Effects of naloxone and morphine on gastric acid secretion and on serum gastrin and pancreatic polypeptide concentrations in humans, Gastroenterology 79, 294. Gascoigne, A.D., B.H. Hirst, J.D. Reed and B. Shaw, 1980, Effects of thyrotropin-releasing hormone and methionine-enkephalin on gastric acid and pepsin secretion in the cat, Br. J. Pharmacol. 69, 527. Holton, P. and J.E. Spencer, 1976, Acid secretion by guinea-pig isolated stomach, J. Physiol. 255,465. Konturek, S.J., J. Tasler, M. Cieszowski, E. Millos, D. Coy and A. Schally, 1980, Comparison of methionine-enkephalin and morphine in the stimulation of gastric acid secretion in the dog, Gastroenterology 78,294.
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© Elsevier/North-Holland Biomedical Press Short communication T H E A C T I O N O F M O R P H I N E A N D N A L O X O N E O N A C I D S E C R E T I O N BY T H E R A T ISOLATED STOMACH s. PAUL CANFIELD 1 and JAN SPENCER 2 1 Department o f Physiology, St. Mary's Hospital Medical School, London W2 1PG, and 2 Department o f Zoology, Westfield College, London NW3 7ST, U.K.
Received 14 November 1980, accepted 19 February 1981
S.P. CANFIELD and J.E. SPENCER, The action o f morphine and naloxone on acid secretion by the rat isolated stomach, European J. Pharmacol. 71 (1981)135--138. The action of morphine and naloxone on acid secretion by the rat isolated stomach has been studied. Morphine (10 -7 to 10 -4 M) had no effect upon spontaneous acid secretion. Morphine (10 -5 M) did not modify the acid output in response to sub-maximal stimulation by pentagastrin, histamine, bethanechol or isoprenaline. Naloxone (10 -5 M) was without effect on the response to pentagastrin or histamine. Our results suggest that opiate receptors do not modify acid secretion in this preparation. Morphine
Rat
Naloxone
Isolated
1. I n t r o d u c t i o n T h e r e have r e c e n t l y b e e n a n u m b e r o f p u b lications c o n c e r n e d w i t h t h e a c t i o n o f o p i a t e agonists o n gastric acid s e c r e t i o n in vivo. K o n t u r e k et al. ( 1 9 8 0 ) r e p o r t e d t h a t b o t h m o r p h i n e a n d M e t - e n k e p h a l i n caused dose d e p e n d e n t increases in acid s e c r e t i o n in b o t h H e i d e n h a i n p o u c h and fistula c o n s c i o u s d o g p r e p a r a t i o n s . T h e s e r e p o n s e s were i n h i b i t e d b y n a l o x o n e a n d also b y a t r o p i n e a n d m e t i a m i d e G a s c o i g n e et al. ( 1 9 8 0 ) f o u n d n o e f f e c t o f M e t - e n k e p h a l i n o n e i t h e r basal or stimulated acid s e c r e t i o n in fistula cats even at d o s e s high e n o u g h to cause a t o x i c r e a c t i o n . E1 Munshid et al. ( 1 9 8 0 ) r e p o r t e d t h a t Mete n k e p h a l i n i n h i b i t e d h i s t a m i n e , b u t n o t pentagastrin, s t i m u l a t e d acid s e c r e t i o n in conscious fistula rats. In h u m a n subjects, F e l d m a n et al. ( 1 9 8 0 ) f o u n d t h a t n a l o x o n e i n h i b i t e d acid s e c r e t i o n f o l l o w i n g a meal. H o w e v e r , m o r p h i n e also caused a r e d u c t i o n o f acid output. T h e s e studies d o n o t p r e s e n t a simple pict u r e o f the e f f e c t o f o p i a t e agonists o n acid
Acid
Stomach
secretion. In p a r t t h e d i f f e r e n c e s in t h e results m a y be d u e s i m p l y to species d i f f e r e n c e b u t m a y also be d u e t o t h e wide s p e c t r u m o f activity o f this class o f drugs in t h e b o d y . Opia t e agonists h a v e b e e n r e p o r t e d to increase gastric m u c o s a l b l o o d flow, cause the release o f h i s t a m i n e a n d gastrin and i n h i b i t the release o f a c e t y l c h o l i n e . T h u s o p i a t e s m a y m o d i f y acid s e c r e t i o n b y a v a r i e t y o f i n d i r e c t m e a n s r a t h e r t h a n b y a p r i m a r y a c t i o n on t h e parietal cell. We have t h e r e f o r e s t u d i e d t h e a c t i o n o f m o r p h i n e a n d n a l o x o n e o n acid s e c r e t i o n using an in vitro s t o m a c h p r e p a r a t i o n f r o m t h e r a t w h e r e s o m e o f t h e s e difficulties are absent.
2. Materials a n d m e t h o d s 2.1. P r e p a r a t i o n
S t o m a c h s were o b t a i n e d f r o m i m m a t u r e Wistar rats weighing 3 0 - - 5 0 g w h i c h h a d b e e n k e p t w i t h a lactating f e m a l e . A n i m a l s w e r e killed b y cervical d i s l o c a t i o n and t h e s t o m a c h
136
S.P. CANFIELD, J. SPENCER
rapidly r e m o v e d , cut along the lesser curvature, everted and washed with cold 0.9% saline solution. The s t o m a c h s were tied over the end o f a p e r s p e x t u b e (cross sectional area 1.1 cm 2) with the m u c o s a facing into the l u m e n o f t h e tube. The m u c o s a was b a t h e d by 5 ml o f u n b u f f e r e d saline gassed with 100% 02 and the serosal surface with 30 ml o f HCO~ b u f f e r e d saline (pH 7.4) gassed with 95% 02-5% CO2. T h e solutions were maintained at 34°C and their c o m p o s i t i o n was identical with t h a t previously used for the guinea pig s t o m a c h b y H o l t o n and S p e n c e r (1976). T h e mucosal saline was replaced at 15 min intervals, its pH m e a s u r e d and it was t i t r a t e d t o pH 7 with 20 mM NaOH.
over a 4 h period. All drugs were a d d e d to the serosal saline and left in c o n t a c t with the tissue for 1 h.
2.3. Drugs All drug dilutions were freshly m a d e on the day o f the e x p e r i m e n t and the following were used: pentagastrin (Peptavlon, ICI Ltd, U.K.), isoprenaline sulphate, histamine acid sulphate (Macarthys L t d , U.K.), b e t h a n e c h o l chloride ( G l e n w o o d L a b o r a t o r i e s Ltd, U.K.), Naloxo n e h y d r o c h l o r i d e (Narcan, W i n t h r o p e , U.K.) and m o r p h i n e sulphate (Evans Medical Ltd, U.K.).
2.4. Statistics 2.2. Expression of results A variable p r o p o r t i o n o f the i m m a t u r e rat s t o m a c h p r e p a r a t i o n consists of n o n - s e c r e t o r y r u m e n and part o f this was included within the area o f the perspex t u b e in some preparations w h e r e t h e r u m e n was relatively large. This r e d u c e d the area of s e c r e t o r y m u c o s a to less t h a n 1.1 cm 2. As it was n o t possible t o estimate a c c u r a t e l y the size o f this r e d u c t i o n t h e responses t o drug a d d i t i o n have been expressed as the s e c r e t o r y ratio (R) w h e r e R=
Average plateau response to drug Average spontaneous secretion
T h e average s p o n t a n e o u s secretion was obt a i n e d f r o m t h e rates for the t w o periods imm e d i a t e l y prior t o drug addition. Thus a value o f R = 1 indicates no stimulation whilst R = 2 shows a d o u b l i n g o f t h e rate o f secretion in t h e presence o f the drug. Thus, the spontaneous secretion (usually 2--4 ~ m o l H ÷ cm -2" h -1) has b e e n utilised as an i n d e x o f the a m o u n t o f s e c r e t o r y mucosa. S o m e absolute m e a n values o f acid secretion (response-spont a n e o u s in p m o l H ÷ cm -2 • h -1) with standard errors have b e e n included in the t e x t to facilit a t e c o m p a r i s o n o f this p r e p a r a t i o n with o t h e r published w o r k on isolated stomachs. C o n t r o l e x p e r i m e n t s s h o w e d t h a t the spont a n e o u s secretion did n o t change significantly
Comparisons b e t w e e n test s t o m a c h s and c o n t r o l s t o m a c h s were m a d e on the s e c r e t o r y ratios using the Mann-Witney U-test as this has the advantage t h a t it does n o t require assumptions a b o u t the statistical distribution o f the data.
3. Results
3.1. Action of morphine alone M o r p h i n e was a d d e d t o t h e s t o m a c h s b y c u m u l a t i v e increases in c o n c e n t r a t i o n over the range 10 -7 to 10 -4 M. T h e m e a n values o f R with standard errors are s h o w n in fig. 1. Each c o n c e n t r a t i o n was left in c o n t a c t for 1 h, t h e n washed o u t and i m m e d i a t e l y replaced by the higher c o n c e n t r a t i o n . N o n e o f the c o n c e n t r a tions used caused any significant change in the s p o n t a n e o u s secretion. T w o p o i n t concent r a t i o n - r e s p o n s e curves f o r histamine and isoprenaline have b e e n included for c o m p a r i s o n . In absolute values the response at 1 0 - 4 M m o r p h i n e was --0.17 + 0.11 p m o l H ÷ c m -2 • h -~, n o t significantly d i f f e r e n t f r o m zero. The responses to the higher c o n c e n t r a t i o n s o f hist a m i n e and isoprenaline were respectively 1.60 _+ 0.29; 3.19 _+ 0.48 p m o l H* cm -2 • h -1.
MORPHINE AND ACID OUTPUT BY RAT ISOLATED STOMACH
2.2 ~13 2.0 1.6 L2
137
TABLE 1 The effect of morphine (10 -6 M) and naloxone (10 -6 M) on secretagogue induced acid secretion in the rat isolated stomach. Test stomachs were preincubated with either morphine or naloxone for 1 h prior to addition of either pentagastrin (P), histamine (H), bethanechol (B) or isoprenaline (I) for a further hour. Control stomachs received only the secretagogues. Mean values with standard error and number of observations in brackets are shown. Probability values were obtained from Mann-Witney U-tests.
1.8
14
,.o ~_________~÷
Secretagogue
°'2,
,o
Drug
Concentration
,00
Secretory test
(microMolar)
Fig. 1. Secretory ratio R as ordinate plotted against log drug concentration as abscissa. Points are mean values with standard error bars. The number of observations (n) was 8 for all morphine points (A A). Two point curves for isoprenaline (e ¢,) and histamine (u --) are included for comparison and n is shown beside each point.
3.2. A c t i o n o f m o r p h i n e on o t h e r secretagogues We also i n v e s t i g a t e d t h e e f f e c t o f preincubating the stomachs with morphine ( 1 0 -6 M) f o r 1 h o n t h e s e c r e t o r y r e s p o n s e t o approximately half-maximal concentrations of either pentagastrin, histamine, bethanechol, or isoprenaline. Results were c o m p a r e d with identically treated control stomachs w h i c h d i d n o t h a v e m o r p h i n e . T h e r e s u l t s are s h o w n i n t a b l e 1A. C l e a r l y m o r p h i n e d i d n o t alter the response compared with the controls. For b e t h a n e c h o l the c o n t r o l secretion w a s 1 . 9 9 _+ 0 . 5 1 ~zmol H ÷ c m - 2 • h -1 a n d i n t h e presence of morphine 2.71 ± 0.86 pmol H ÷ c m - 2 . h-1.
3.3. A c t i o n o f n a l o x o n e Konturek e t al. ( 1 9 8 0 ) r e p o r t e d t h a t naloxone inhibited sub-maximal responses to p e n t a g a s t r i n a n d h i s t a m i n e as well as i n h i b i t ing o p i a t e - s t i m u l a t e d secretion in their cons c i o u s d o g s t u d i e s . We t h e r e f o r e e x a m i n e d the effect of naloxone (10-6M) on the
Ratio (R) control
Probability
0.24
A. Morphine (10 -6 M) P (10 -7 M)
1.65 (6) + 0.06
1.60 (6) + 0.09
H(4.4 × 1 0 - s M )
1.59 (8) + 0.04
1.47 (9) + 0.05
B (8 × 10 -6 M)
2.22 (6) + 0.31
2.02 (6) +- 0.39
0.35
I (6 × 10 -7 M)
1.79 (6) -+ 0.10
1.75 (6) + 0.12
0.16
1.48 (6) + 0.11
0.53
1.41 (6)
1.40 (6)
0.41
± 0.03
± 0.06
>0.1
B. Naloxone (10 -6 M) P (10 -7 M) H (4.4 × 10 -s M)
1.53 (5) + 0.10
response of the rat stomach to half-maximal concentrations of histamine and pentagastrin u s i n g t h e s a m e t e s t - c o n t r o l d e s i g n as a b o v e . T h e r e s u l t s are s h o w n i n t a b l e l B . N a l o x o n e had n o effect o n the responses at this c o n c e n tration. The absolute values for pentagastrin w e r e 1 . 9 0 + 0 . 2 7 p m o l H ÷ c m - : • h -1 f o r c o n t r o l s a n d 1 . 5 5 + 0 . 3 9 p m o l H * c m -2 . h -1 with naloxone. The spontaneous secretion for the p e n t a g a s t r i n c o n t r o l s was 3 . 1 6 + 0.48 p m o l H ÷ c m -2 • h -1 w h i l s t i n t h e p r e s e n c e o f n a l o x o n e it was 3 . 6 5 +_ 0 . 9 6 ~ m o l H + c m -2 • h -1. T h e y w e r e n o t s i g n i f i c a n t l y d i f f e r e n t .
4. D i s c u s s i o n Morphine did not have any effect on either s p o n t a n e o u s acid secretion or t h a t s t i m u l a t e d
138
by any of the four secretagogues used and naloxone was also without action on the two stimulants tested. In presenting negative results there is always the problem that they may arise from the use of poor preparations rather than indicating the behaviour of the system being studied. The stomachs used in this study secreted acid spontaneously and responded to four different stimulants. Although the number of observations was not large we have previously shown inhibitory effects in this preparation using other drugs in similar sized samples (Canfield et al., 1980). The use of secretory ratios rather than absolute values for the statistical tests would tend to favour positive results as the within treatment variance of the ratios tends to be lower than that with absolute values where no allowance is made for the variable a m o u n t of secretory mucosa. The secretory ratio might conceal changes in response if the spontaneous secretion were altered by the drug treatment under test in such a way that the ratio derived from this was unaltered. However, neither morphine nor naloxone altered spontaneous acid output. We therefore, feel confident that our negative results with morphine and naloxone indicate that there are no opiate receptors in the
S.P. CANFIELD, J. SPENCER
rat stomachs capable of modifying acid secretion. There remains the possibility that opiate receptors outside the stomach may influence acid secretion indirectly.
References Canfield, S.P., C.A. Price and J.E. Spencer, 1980, Noradrenaline and gastric acid secretion by the rat isolated stomach, Br. J. Pharmacol. 70,178P. El Munshid, H.A., R. Hakanson, G. Liedberg and F. Sundler, 1980, Effects of various gastrointestinal peptides on parietal cells and endocrine cells in the oxyntic mucosa of rat stomach, J. Physiol. 305,249. Feldman, M., J.H. Walsh and L.L. Taylor, 1980, Effects of naloxone and morphine on gastric acid secretion and on serum gastrin and pancreatic polypeptide concentrations in humans, Gastroenterology 79, 294. Gascoigne, A.D., B.H. Hirst, J.D. Reed and B. Shaw, 1980, Effects of thyrotropin-releasing hormone and methionine-enkephalin on gastric acid and pepsin secretion in the cat, Br. J. Pharmacol. 69, 527. Holton, P. and J.E. Spencer, 1976, Acid secretion by guinea-pig isolated stomach, J. Physiol. 255,465. Konturek, S.J., J. Tasler, M. Cieszowski, E. Millos, D. Coy and A. Schally, 1980, Comparison of methionine-enkephalin and morphine in the stimulation of gastric acid secretion in the dog, Gastroenterology 78,294.