The action of choline on the superior cervical ganglion of the cat

EUROPEAN JOURNAL OF PHARMACOLOGY 17 (1972) 87-96. NORTH-HOLLAND PUBLISItlNG COMPANY

THE ACTION OF CHOLINE

ON THE SUPERIOR

CERVICAL

GANGLION

OF THE CAT *t

M.K. K R S T I ( Department of Pharmacology, Medical Faculty, Beograd 11105, Yugoslavia

Received 29 September 1970

Accepted 23 September 1971

M.K. KRSTIC, Th e action of choline on the superior cervical ganglion of the cat, European J. Pharmacol. 17 (1972) 87 -96. General characteristics and pharmacological properties of the action of choline on the superior cervical ganglion (SCG) were studied and compared with those of dimethylphenylpiperazinium (DMPP). Chronic denervation of the SCG decreased, while preganglionic stimulation (single supramaximal and a series of intermittent submaximal) potentiated the response of this ganglion to choline. Hexamethonium blocked, while atropine considerably reduced the ganglionic effect of choline. The effect of choline on the SCG was blocked by small doses of nicotine; it was strongly reduced but not blocked by large doses. Cocaine (particularly in adrenalectomized cats), morphine and methadone considerably reduced or blocked the ganglionic effect of choline. It is concluded that there is an atropine-sensitive component in the action of choline on the SCG. The hypothesis is proposed that within the SCG there is either an intermediate type of receptor having some properties of both nicotinic and atropine-sensitive receptors or that the specificity of the latter receptors is more limited than is presently thought. Superior cervical ganglion (SCG) Ganglionic effect

Nictitating membrane (NM) Contractions of the NM

INTRODUCTION Dale (1914) has demonstrated that choline causes hypertension in the atropinized cat which can be blocked by large doses o f nicotine. Small doses o f choline stimulate, while large ones paralyze the perfused: superior cervical ganglion (SCG) of the cat (Feldberg and Vartiainen, 1934). Gebber and Voile (1965) have shown that choline, injected i.a., causes deplorization o f the SCG and postganglionic firing. Hexamethonium blocks both o f these effects o f choline. The same authors, by conditioning the SCG with repetitive preganglionic stimulation, anticholinesterase agents or isoproterenol, were able to unmask a late-occurring atropine-sensitive component in the postganglionic response to choline. In the present ex* A preliminary report of the results was presented at the Vlth Congress of the Yugoslav Physiological Society, September 1969, Ohrid, Yugoslavia. t This work was supported by Union of the Scientific Medical Institutions (Grant No. 16).

Dimethylphenylpiperazinium (DMPP) Choline

periments an attempt was made to detect, without prior conditioning of the ganglion, the atropine-sensitive component in the response o f the SCG to choline. A study on the general characteristics o f the effect of choline of the SCG preceded these experiments.

2. MATERIALS AND METHODS Cats o f b o t h sexes, weighing 1 . 7 - 3.4 kg, were anaesthetized with a mixture o f chloralose, 6 4 80 mg/kg, and urethane, 4 0 0 - 5 0 0 mg/kg, given i.v. The trachea was cannulated. Arterial blood pressure was recorded directly from the femoral artery using a mercury manometer and a smoked drum. Before the experiment was started, about 1000 I.U. o f heparin was injected into the cannula inserted into the femoral artery. Throughout each experiment the rectal temperature was kept at about 36°C. The effect o f choline on the SCG was studied by

88

M.K. Krsti(, Choline on the superior cervical ganglion

the method of Trendelenburg (1959). 1.a. injections were made through a polyethylene cannula inserted into the right lingual artery. Drugs injected into this cannula could be directed towards the SCG ('towards the ganglion') by occluding the external carotid artery. The drug could be directed towards the nictitating membrane ('towards the membrane') by leaving the external carotid artery unoccluded. The contractions of the nictitating membrane (NM)were magnified 12 times and recorded by an isotonic frontal lever on a smoked drum. In order to prevent drying of the NM it was covered with a thin layer of paraffin oil. In the majority of experiments the effect of choline on the SCG was analysed immediately after cutting the cervical sympathetic chain low in the neck. However, in one experimental group the right SCG was denervated, i.e. the right NM was decentralized. Cats in this g r o u p were anaesthetized intraperitoneally with pentobarbital-sodJum, 2 5 - 30 mg/kg, and, by an aseptic operation, a 1 cm section o f the cervical sympathetic chain was removed. Penicillin was applied locally to the wound and was also injected intramuscularly for 3 days. These cats were used for experiments 9 - 14 days after the cutting o f the cervical sympathetic chain. In these animals polyethylene cannulae were inserted into both lingual arteries, so that the effect of choline on the normal and the denervated SCG could be examined in the same cat. In some acute experiments the right SCG was removed surgically. In other acute experiments, cats were bilaterally adrenalectomized transabdominally after carefully tying the vessels o f the suprarenal glands. For electrical stimulation of the cervical sympathetic chain, it was cut a n d its peripheral end was placed on shielded platinum electrodes and covered with liquid paraffin. Electrical stimulation was applied with a stimulator delivering rectangular impulses of 0.75 msec duration at a frequency of 25/sec. Either intermittent submaximal preganglionic stimulation for 5 sec every 30 sec or one 30 sec period of supramaximal preganglionic stimulation was used. The ganglionic effect of DMPP was used as a control. In each experiment the time intervals between the injections of choline towards the ganglion, as well as between the injection of choline and DMPP were approximately the same. Drugs were dissolved in isotonic NaC1 solution and

injected i.a. in a volume of 0.1 0.2 ml. Tile circulation in the external carotid artery was stopped by a clamp 0.5 - 1 rain before recording the ganglionic effects of choline or DMPP. The circulation was reestablished after the drug-induced contraction o f the NM was complete and after removal of any drug solution which might have remained in the common carotid artery. The latter was accomplished by allowing a few drops of blood to flow back from the free end of the cannula, which was then refilled with isotonic NaCI solution. In each experiment, choline and DMPP were injected i.a. at the same rate (2 or 3 sec depending on the volume of the solution). Drugs were injected i.v. through a polyethylene cannula inserted into the left femoral vein. The following substances were used: choline chloride; dimethylphenylpiperazinium iodide (DMPP), adrenaline hydrochloride; synthetic bradykinin (BRS 640, Sandoz), hexamethonium bromide, nicotine hydrogen tartarate, atropine sulphate, morphine hydrochloride, methadone hydrochloride (Heptanon, Pliva) and cocaine hydrochloride. All doses refer to the salts. 3. RESULTS 3.1. The effect o f choline Choline, 5 0 - 6 0 0 # g , injected towards the ganglion, after a latent peri.od from 1.5 and 5 sec, caused 3

C

O • • • • • rn O • • Fig. 1. Contractions of the NM of the cat (3 kg). The effect of submaximal intermittent preganglionic stimulation (SIPS) on the response of the SCG to choline. At dots, 100 tag of choline was injected towards the ganglion; at square, 100 tag of choline towards the membrane; and at circles, 2 tag of adrenaline towards the membrane. Between B and C one series of 8 single SIPS for 5 sec, at a frequency of 25/sec, every 30 sec. Time interval between A and B 4.5 min, B and C 13 min, and C and D 80 min.

M.K. Krsti6, Choline on the superior cervical ganglion

:~

3.2. The effect o f choline compared with that o f DMPP

60

Contractions of the NM produced by the injection of choline towards the ganglion were similar in shape to contractions of the NM produced by DMPP. There was, however, a difference in the speed and duration of these contractions (table 1). On a weight basis, the effect of choline on the SCG is 1 0 0 - 2 0 0 times weaker than the effect of DMPP.

50 "5

.~ E 30 oLJ

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10 0 200

dose in j u g / k g ([og.scole)

Fig. 2. Relationship between doses of choline and height of the contractions of the NM. Choline was injected towards the ganglion at a constant concentration (l mg/ml) and rate of injection, at intervals of 15 rain. Every point represents mean of 5 experiments _+S.D.

a contraction of the mictitating membrane (NM) lasting 0.5 - 1.5 min (fig. 1). There was a linear relationship between the logarithm of the doses of choline and the height of the contractions of the NM (fig. 2). On the other hand, when the same dose of choline (120 6 0 0 p g ) was injected at the same rate in different volumes, a linear relationship between the logarithm of the drug concentrations and the height of the contractions of the NM was also found. The height of the contractions did not change appreciably after 7 - 10 injections of the same dose of choline towards the ganglion at intervals of 3 6 min. In 16 out of 30 experiments, however, an increase in the magnitude of the contractions was observed when the same dose of choline was injected at intervals of 14 3 0 m i n . Choline injected towards the membrane did not bring the NM into contraction (fig. 1). Table 1 Latency, speed and duration of the contractions of the NM produced by the injection of choline, 100-300 #g, and DMPP, 1-2 #g, towards the ganglion. DMPP Latent period between injection and beginning contraction 3.3-+0.26 Rising time 7.7-+0.56 Duration contraction 41.0-+1.84

Choline

p value difference

5.4_+0.38 13.5+0.82 63.0-+2.75

< 0.001 < 0.001 < 0.001

Time in sec, means +- S.E.M. from 13 expts. The contractions of the NM produced by DMPP and choline were of approximately the same height.

3.3. Adrenalectomy, ganglionectomy and cutting o f the cervical sympathetic chain There is the possibility that choline injected towards the ganglion partly passes into the general circulation. In this case, contractions of the NM, after administration of choline, could be partially or completely due to the release of catecholamines from the adrenal medulla induced by choline. In order to exclude such a possibility, in one experimental group bilateral adrenalectomy was performed. The contractions of the NM caused by injections of choline towards the ganglion were not decreased after adrenalectomy (8 experiments). Furthermore, the ganglionic origin of these contractions can be proved if they are abolished after removal of the SCG. In 4 separate experiments, choline iniected into the lingual artery during the occlusion of the external carotid artery, after the surgical removal of the SCG, did not produce a contraction of the NM. Since, in the majority of cats, the cervical sympathetic chain was cut just before the experiment, it was of interest to examine the influence of this procedure on the response of SCG to choline. Cutting of the cervical sympathetic chain did not affect the magnitude of the contractions of the NM caused by choline (4 experiments).

3.4. Denervation o f the SCG Lewis and Reit (1965) have shown that the response of the SCG to bradykinin is unchanged after its denervation. The supersensitivity of the decentralized NM to the action of drugs is well known. In one experimental group the levers recording the contractions of the decentralized and the normal NM were equally loaded. When under this condition adrenaline was injected i.v. the contraction of the decentralized NM was 2 - 3 times stronger than that of the normal NM. Under the same conditions, the response of the

90

M.K.Krstik, Choline on the superior cervicalganglion

decentralized NM to the injection of the bradykinin towards the denervated ganglion was also much greater than the response of the normal NM. This observation can be explained on the assumption that bradykinin about equally stimulates the innervated and the denervated postsynaptic neurones (Lewis and Reit, 1965) and that the ensuing liberation of noradrenaline from the nerve endings in the NM produces an increased contraction of the decentralized NM because of its supersensitivity to noradrenaline resulting from the decentralization. Surprisingly, however, no difference was found between the responses of the normal and the decentralized NM to the injection of either choline or DMPP towards the normal and the denervated ganglion. This might be explained on the assumption that the supersensitivity of the decentralized NM was compensated for by a corresponding decrease in the sensitivity of the denervated postsynaptic ganglionic neurones to choline and DMPP. In a second experimental group, the influence of the supersensitivty of the decentralized NM to noradrenaline was excluded by loading the lever recording the contractions of the decentralized NM with less weight than the lever recording the contractions of the normal NM. Under these experimental conditions, the contractions of both NM produced by intravenous injection of adrenaline were of the same height. There were no appreciable differences between the height of contractions of the NM caused by bradykinin injected towards the denervated and towards the normal SCG. However, contractions of the NM caused by injections of choline and DMPP towards the denervated SCG were considerably smaller than the contractions produced by injections of these drugs towards the normal SCG. This supports the idea that the sensitivity of postsynaptic receptor sites in the SCG to choline and DMPP is decreased after denervation. 3.5 Action o f preganglionic stimulation on the ganglionic effect o f choline Contractions of the NM, produced by injection of choline towards the ganglion, 5 min after supramaximal preganglionic stimulation (SMPS), as well as 8 - 12 rain after a series of 8 submaximal intermittent preganglionic stimulations (SIPS) at intervals of 30 sec, were potentiated in height by 1.4 - 4 times (8 experiments). This potentiation of the contractions

of the NM lasted longer than 90 min (fig. 1). Under the same experimental conditions, SMPS and SIPS did not potentiate contractions of the NM caused by injection of adrenaline towards the membrane. Thus, the potentiation of the effect of choline by SMPS and SIPS is of ganglionic origin.

3.6. Effect o f choline on the response o f the N M to submaximal intermittent preganglionic stimulation

(sips) In 5 out of 8 experiments choline, 25 - 200 pg, injected towards the ganglion, reduced or blocked the response of the NM to SIPS, while in 3 experiments it did not change or potentiate the response of the NM. The depression of the response of the NM to SIPS partially or completely passed off 15 - 20 rain after injection of choline. The block was produced by doses of choline which were insufficient to cause a contraction of the NM, as well as by higher doses which were able to elicit a contraction. Choline injected towards the NM did not reduce the contractions of this membrane caused by SIPS. Accordingly, the decrease or abolition of the response of the NM to SIPS results from the depression of synaptic transmission in the SCG induced by choline. 3.7. Effect o f hexamethonium Hexamethonium, 300 - 500 pg, injected towards the ganglion, blocked the stimulant effect of choline and DMPP on the SCG. The effects of both ganglionic stimulants were completely re-established 1 6 - 5 0 min after the injection of hexamethonium. Smaller doses of hexamethonium ( 1 5 0 - 300 #g injected towards the ganglion 5 or 9 rain before recording the ganglionic effect of choline) in 6 out of 14 experiments decreased equally the response of the SCG to choline and DMPP. In the remaining cats, the ganglionic effect of either choline (5 experiments) or DMPP (3 experiments) was more decreased. 3.8. Effect o f atropine Atropine (1 - 1.6 pg injected towards the ganglion 9 - 11 min before recording the effect choline) decreased the ganglionic effect of choline by 50 - 65%, while it did not significantly change the effect of DMPP (5 experiments). The effect of choline on the SCG began to recover 27 - 55 min after application of atropine.

M.K.Krsti6, Choline on the superior cervicalganglion 3.9. Effect o f atropine after administration o f hexamethonium and vice versa In 5 experiments, atropine was injected towards the ganglion when both the ganglionic effect of choline and DMPP had completely recovered from a strong reduction produced by previous administration of hexamethonium. In 2 experiments atropine, 1 - 1.5/ag, blocked and in 3 experiments diminished the ganglionic effect of choline by 60 - 84%, while it did not affect or only slightly decreased the ganglionic effect of DMPP (fig. 3). In 3 experiments, hexamethonium was injected towards the ganglion when the ganglionic effect of choline had completely recovered from a considerable depression caused by previous administration of atropine. Hexamethonium, 100 - 150/ag, blocked the responses of SCG to choline and DMPP in all experiments (fig. 4). The ganglionic effects of choline and DMPP recovered simultaneously. The block of the C

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Fig. 4. Contractions of the NM of the cat (1.7 kg). The effect of atropine and hexamethonium on the response of the SCG to choline and DMPP. At cirles, 1.3 ug of DMPP was injected towards the ganglion; at dots, 150 t~g of choline; between A and B, 1.6 #g of atropine; and between E and F, 100 ~tg of hexamethonium. Time interval between A and B 8 min, B and C 40 min, C and D 50 min, D and E 30 rain, E and F 10 min, and F and G 20 min. gaLglionic effect of choline by hexamethonium and reduction or block by atropine suggest that hexam e t h o n i u m and atropine act on the same type of ganglionic receptors.

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Fig. 3. Contractions of the NM of the cat (2.2 kg). The effect of hexamethonium and atropine on the response of the SCG to choline and DMPP. At dots, 260 ug of choline was injected towards the ganglion; at circles, 2.25 ug of DMPP; between A and B, 300 t~g of hexamethonium; and between D and E, 1.5 ug of atropine. Time interval between A and B 8 min, B and C 10rain, C andD 8min, D and E 10min, E and F 30 min, F and G 20 min, and G and H 21 min.

Small doses of nicotine (100/ag, 3 times or 200/~g twice, at intervals of 90 sec), injected towards the ganglion, blocked the effect of choline on the SCG (fig. 5). The ganglionic effect of choline was completely re-established 15 to 25 min after the administration of nicotine (10 experiments). In 6 experiments, nicotine (total 3.7 mg) was injected towards the ganglion in a series of increasing doses at time intervals of 90 sec. Large doses of nicotine in one experiment blocked, while in 5 experiments only considerably reduced ( 6 5 - 85%), contractions of the NM produced by choline. Fig. 5 shows the experiment in which nicotine produced the greatest diminution of the ganglionic effect of choline. The part of the contractions of the NM which was resistant to large doses of nicotine appeared after an appreciably longer latency, developed more slowly and lasted longer than the control contractions.

M.K. Krsti6, Choline on the superior cervical ganglion

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3.11 Effect of methadone and morphine Methadone, 2 5 - 75/2g/kg, i.v., with a delay of 6 - 20 min after its administration, considerably diminished the ganglionic effect of choline in 6 out of 9 experiments. Morphine, 10 - 30/ag/kg, in 2 out of 4 experiments blocked, and in 2 experiments considerably reduced the ganglionic effect of choline. In both experimental groups the ganglionic effect of DMPP was unchanged or appreciably less affected than the effect of choline (fig. 6). The response of the SCG to choline recovered, in all experiments, 40 - 90 rain after the injection either of methadone or morphine.

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3.12 Effect of cocaine Cocaine (0.45 - 0.7 mg/kg, injected i.v., 5 rain before recording the effect of choline on the SCG) in 4 out of 5 experiments decreased, and in one experiment potentiated contractions of the NM caused by choline. The decrease of the contractions lasted 1 0 18 rain and varied from slight to considerable. In all experiments, however, 20 - 30 min after the administration of cocaine, the effect of choline on the SCG was potentiated. In a separate group of 5 experiments, the action of cocaine, 2 . 0 - 2.3 mg/kg, i.v., on the ganglionic response to choline was examined in adrenalectomized cats. 5 - 20 min after the administration of cocaine, the ganglionic effect of choline was diminished

X

Fig. 5. Contractions of the NM of the cat (3.4 kg). The effect of nicotine on the response of the SCG to choline. At dots, 225 pg of choline was injected towards the ganglion; at arrows, 100 tag of nicotine; and at X, 0.15 ml o f the isotonic solution of NaC1. Between B and C, at time intervals of 90 sec, 5 injections of 200 ug of nicotine, 3 injections of 400 tag of nicotine and 3 injections o f 500 tag o f nicotine were made i.a. towards the ganglion. Time interval between A and B 30 rain, B and C 25 min, C and D 10 min, and D and E 4 min.

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Fig. 6. Contractions of the NM of the cat (1.7 kg). The effect of morphine on the response of the SCG to choline and DMPP. At dots, 200 pg of choline was injected towards the ganglion; at circles, 1.5 #g o f DMPP towards the ganglion; between A and B, 27 tag of morphine i.v.; and between D and E, 60 tag o f m o r p h i n e i.v. Time interval between A and B 9 min, B and C 7 min, C and D 7 rain, D and E 14 min, and E and F 50 min.

M.K.Krsti6, Choline on the superior cervical ganglion

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Fig. 7. Contractions of the NM of the bilaterally adrenalectomized cat (2.6 kg). The effect of cocaine on the response of the SCG to choline and DMPP. At dots, 150 #g of choline was injected towards the ganglion;at circles, 1.5 t~g of DMPP towards the ganglion; and between A and B, 6 mg of cocaine i.v. Time interval between A and B 15 min, and B and C 50 min.

( 3 5 - 55%) in all experiments, while the effect of DMPP was potentiated in height as well as in the duration (fig. 7). The reduction of the ganglionic effect of choline produced by cocaine lasted longer than 50 rain. Thus, the appreciable depressant action of cocaine on the ganglionic effect of choline was evident only after bilateral adrenalectomy of the cat.

4. DISCUSSION Choline, injected towards the ganglion, contracted the NM. The same dose of choline injected towards the membrane, however, did not produce a contraction. Furthermore, the ganglionic origin of the contractions of the NM is confirmed by their abolition after surgical removal of the SCG. Trendelenburg (1956a) has found that choline depresses transmission of nerve impulses through the perfused SCG of the cat. In the present experiments, it has been demonstrated that choline can also depress synaptic transmission in the SCG with intact blood supply. Gebber and Voile (1965) have shown that supramaximal preganglionic stimulation enhances the postganglionic firing produced by choline. Our results support this finding. Furthermore, the present experi-

93

ments show that the response of SCG to the effect of choline is also potentiated after a series of submaximal intermittent preganglionic stimulations. The characteristics of the lattor potentiation are similar to those of the former. In the present experiments the responses of the denervated SCG to the action of choline and DMPP were considerably decreased. However, the response of the denervated SCG to the action of bradykinin was not appreciably changed. These findings could be explained: (1) by a stimulating action of choline and DMPP, in part, on presynaptic nerve terminals in the SCG, (2) by a decrease of the sensitivity of postsynaptic receptor sites in the denervated SCG and (3) by the presence of both events. It is known that bradykinin activates only postsynaptic receptor sites in the SCG (Lewis and Reit, 1965). According to present concepts, denervation of the SCG does not change the sensitivity of the nicotinic receptors, while it increases greatly that of the atropine-sensitive receptors (Trendelenburg, 1967). The potentiation of the ganglionic effect of choline by supramaximal and intermittent submaximal preganglionic stimulation shows some resemblance to post-tetanic potentiation of ganglionic transmission, which is a presynaptic event (Voile, 1966). Moreover, according to some authors (Riker and Szreniawski, 1959; Volle and Koelle, 1961; McKinstry et al., 1963; McKinstry, 1965; McKinstry and Koelle, (1967) acetylcholine and carbachol act on both presynaptic nerve endings and postsynaptic receptor sites in the SCG. These data speak in favor of the possibility that choline and DMPP activate presynaptic nerve endings of the SCG in addition to postsynaptic receptor sites. However, some recent findings deny the activation of presynaptic nerve endings in the SCG by acetylcholine and carbachol (Brimblecombe and Sutton, 1968; Brown, 1966, 1969; Brown et al., 1970). This supports the possibility that the sensitivity of postsynaptic receptor sites of the denervated SCG to choline and DMPP is decreased. Thus, on the basis of our present knowledge, it is not possible to say which one of the three alternative explanations of the decrease of the denervated SCG response to choline and DMPP is correct. DMPP is a typical nicotine-like ganglionic stimulating agent (Ling, 1955; Gumulka and Szreniawski; 1968) Hexamethonium, depending on the injected dose, blocked or equally reduced the ganglionic ef-

94

M.K.Krstid, Choline on the superior cervical ganglion

fects of choline and DMPP. However, atropine considerably reduced the effect of choline on the SCG, while the effect of DMPP was unchanged or slightly decreased. Gebber and Voile (1965), after conditioning of the SCG by repetitive preganglionic stimulation, anticholinesterase agents or isoproterenol, discovered a late-occurring atropine-sensitive component in the postganglionic response to choline. The present experiments demonstrate this atropine-sensitive component without prior conditioning of the ganglion. The contractions of the NM caused by injections of nicotinic substances (DMPP, acetylcholine, tetramethylammonium and nicotine) towards the ganglion differ from the contractions caused by injections of muscarinic substances (muscarine, substance McN-A343 and substance AHR-602) in latent period, in speed and in duration (Jones, 1963). The contractions of the NM caused by choline began after a longer latent period, developed slower and lasted longer than the contractions produced by DMPP. This finding also distinguishes the ganglionic effect of choline from the nicotine-like ganglionic stimulants. Morphine, methadone and cocaine reduce or block the effect of non-nicotine-like ganglionic stimulants without changing the effect of nicotine-like ganglionic stimulants (Trendelenburg, 1954, 1955, 1956b, 1957a, 1961, 1966; Jones, 1963; Jones et al., 1963; Murayama and Unna, 1963). In the present experiments morphine and methadone, injected in doses which did not decrease or slightly decrease the ganglionic effect of DMPP, blocked or considerably reduced the ganglionic effect of choline. Cocaine reduced the effect of choline on the SCG, whereas it potentiated the effect of DMPP. The depressant actions of morphine, methadone and cocaine on the ganglionic effect of choline prove its non-nicotine-like characteristics. The depressant effect of cocaine on the NM contractions elicited by choline, injected towards the ganglion, was much more apparent in bilaterally adrenalectomized than in normal animals. Cocaine is a potent antagonist to the effects of the non-nicotine-like agents on the SCG, while it fails to antagonize the effects of these agents on the adrenal medulla of the cat (Slater and Dresel, 1952; Trendelenburg, 1954, 1961, Jones et al., 1963; Franko et al., 1963). It can be supposed that, after an injection of choline towards the ganglion, a small portion of the drug passes into the general circulation, producing the re-

lease of catecholamines from the adrenal medulla. The amount of these catecholamines compared with that released from nerve endings in the NM, due to activation of the SCG by choline, is insignificant and has no effect (see experiments with adrenalectomy). However, after the treatment of the cat by cocaine and the ensuing development of the well-known supersensitivity of the NM, catecholamines released from the adrenal medulla become effective. It is highly probable that, in the presence of such supersensitivity of the NM, the action of these catecholamines can appreciably compensate for the antagonistic effect of cocaine on the SCG response to choline. Nicotine blocks the effects of nicotine-like as well as non-nicotine-like ganglionic stimulants during the deloparizing phase of the block of the SCG, while in the non-depolarizing phase of the block only the effects of nicotine-like ganglionic stimulants are aboli ished (Trendelenburg, 1957b, 1961; Jones, 1963;: Jones et al., 1963). The large doses of nicotine strongly reduced, but did not completely block, the stimulant effect of choline on the SCG. Furthermore, the part of ganglionic effect of choline which was resistant to large doses of nicotine had clearly non-nicotine-like characteristics: long latency and long duration. The effect of hexamethonium, atropine, morphine, methadone, cocaine and nicotine on the response of SCG to choline indicate that this drug has some characteristics of ganglionic stimulants which activate the nicotinic receptors and some characteristics of ganglionic stimulants which activate the atropine-sensitive receptors of the sympathetic ganglia. Atropine considerably reduced the ganglionic effect of choline, and could block it after pretreatment of the SCG with hexamethonium. However, hexamethonium by itself blocked the stimulant effect of choline on the SCG. If choline at the same time activates both nicotinic and atropine-sensitive receptors of the SCG, hexamethonium would be expected to reduce, but not block, the ganglionic effect of this substance. Brimblecombe and Sutton (1968) have shown that hexamethonium blocks the stimulant effect of some amino-acid esters on the SCG, while atropine reduces and sometimes blocks this effect. According to these authors the amino-acid esters cause ganglionic stimulation by interaction with both

M. K. Krsti6, Chbline on the superior cervical ganglion nicotinic and atropine-sensitive receptors in the ganglion. It was concluded that the complete block of the ganglionic effect of the amino-acid esters with hexamethonium could be explained by the property of this ganglionic blocking agent to reduce the ganglionic stimulating effect of 'muscarinic' (non-nicotine-like) drugs. However, many other authors have shown that hexamethonium does not diminish the effect of non.nicotine-like ganglionic stimulants (Root, 1951; Trendelenburg, 1954, 1955, 1961; Roszkowski, 1961; Pappano and Voile, 1962; Levy and Ahlquist, 1962: Jones et al., 1963; Jones, 1963; Murayama and Unna, 1963; Frank et al., 1963; Gokhale et al., 1964; Krsti6, 1968). The properties of the stimulant effect of choline and the amino-acid esters suggest the possibility that within the SCG there exists an intermediate type of receptor having some of the pharmacological characteristics of both nicotinic and atropine-sensitive receptors. If this is not the case, it must be concluded that the specificity of nicotinic and atropine-sensitive receptors in the SCG is more limited than is presently thought.

ACKNOWLEDGMENT The author is indebted to Professor V. Varagi6 for many useful discussions and suggestions, to Dr. T. Rudy for his editorial assistance, and to Mr. Graham Chen from Parke, Davis and Company for supplying dimethylphenylpiperazinium.

REFERENCES

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