t Brit.J,Pharmacol. 49, 706-708 (1973).

706

Short cor_tmlotications 2 hours. The tissue was finally suspended in a 3 ml organ bath maintained at 36°C Inhibition of noradrenaline and gassed with 95 O,/oO,,,: 5%CO,.. The intramural nerves were excited by a field release by lysergie acid current (1 ms rectilinear pulses of supra- diethylamide maximal strength) applied between platinum electrodes set in the top and J. HUGHES bottom of ,the bath. Contractions were , monitored with an isometric transducer Department of Pharmacology, coupled to a Grass Polygraph. To study University o] Aberdeen, Scotland transmitter release the vas was stimulated at 0-2 Hz for 3"5 rain every 15 minutes. .... Lysergicacid diethylamide(LSD)inhibits the The bath fluid was removed 30 s after the release of'labelled noradrcnaline from the end of each stimulation period and the guinea-pig vas deferens during intramural labelled catecholamines were isolated by nerve stimulation and causes a corresponding alumina column chromatography (Boadle- reduction in the contractions of the smooth Biber, Hughes & Roth, 1970). The activity of the alumina eluate was determined in muscle. These effects of LSD are most pro- minent at low stimulus frequencies and they a Packard liquid scintillation counter ,_ are prevented by treatment with phentolamine. (Model 2425). Basal samples for the 4 rain _ It is concluded that LSD inhibits noradren- preceding the onset of each stimulation aline release by interacting with presynaptic period were taken and treated as described above, the basal radioactivity was deducted c_-adrenoceptors, from the stimulated efflux. Attention has focussed mainly on two aspects of the pharmacology of lysergic Results.--LSD tartrate caused a rapid acid diethylamide (LSD), firstly its ability and prolonged inhibition of the contrac- .... to antagonize the effects of 5-hydroxy- tions to electrical stimulation. The effect tryptamine at peripheral (Gaddum, 1957) was most marked at low stimulus frequen- and central sites (Boakes, Bradley, Briggs cies (0.I to 0.5 Hz), and at frequencies & Dray, 1970), and secondly the possibility above 5 Hz was apparent only for the that it may actually stimulate central first 5 to 15 pulses. A 50% inhibition of tryptaminergic receptors by mimicking the the twitch height at 0-1 Hz was obtained action of endogenous 5-hydroxytryptamine with a concentration of 18_+4 ng/ml (Anddn, Corrodi, Fuxe & H6kfelt, 1968}. (n=4), and complete inhibition with 50- I wish .to report a further pharmacological 100 ng/ml (2 rain contact time). The T action of LSD, namely the ability to for recovery of the twitch at 0-1 Hz after- inhibit noradrenaline release from adren- complete inhibition was 22+5 minutes. ergic motor nerves in the guinea-pig was Contractions of the vas deferens elicited ' deferens. This work was prornpted by the by noradrenaline were either unaffected observation that contractions of the vas or slightly potentiated by LSD. Noradrena- 'i deferens elicited by intramural nerve line itself (0"1-1.0 #g/ml) first potentiated | stimulation were reduced by LSD and then depressed the response to elec- IAmbache, Dunk, Miall & Aboo Zar, trical stimulation at 0.I Hz. 1971), although the authors did not con- Electrical stimulation of the vas for sider this effect to be mediated via an 3.5 rain at 0"2 Hz caused a reproducible adrenergic mechanism, increase in the efflux of [3H]-eatechol- amines above basal levels (Figure IL The Methods.--Isolated preparations of the basal efflux of label was unaffected by desheathed vas deferens were incubated LSD (50 ng/ml) but the stimulation eMux in Krebs solution at 36°C with either was reduced to 20-40% of the pre-drug (--)-[aH]-noradrenaline (100 ng/ml, 40 wdue (Figure 1). The contractions and the /_Ci/#g; Amersham Radiochemicals) or stimulated efflux gradually recovered after L-3,5-[_H]-tyrosine (0,25 /.tg/ml, 221 izCi/ washing out the LSD and reintroduction i_g; Amersham Radiochemicals) for 1 or of the drug caused a further ]nhibiti0n of 2 h respectively. At the end of the incuba- the contraction and efftux. The same result tion period the vas was rinsed in fresh was obtained irrespective of whether [_H]- Krebs solution (changed every 10 rain) for noradrenaline or ['_H]-tyrosine was used to Short communications 707

l ! .... t'=_D LSD I d/rai(nx I0-_)I '*="-' ==="=' 5 ! 4"

2

.G' " _-Phentolamine- •_. treated Fl_. l. Antagonism of lysergic acid diethylamide (LSD) by phentolamlne. Vas deferens labelled with [aH]-noradrenaline and field-stimulated as described in Methods. The upper part of the diagram shows the tension developed in the vas3 during periods of stimulation• The lower part of the diagram shows the increased levels of H-labelled material released into the bath during periods of stimulation in the absence (hatched columns) and in the presence (filled columns) of LSD (50 ng/ml). For the experiments recorded on the right-hand side, phentolamine (2 _g/ml) xvas present in the bathing solution.

label the tissue stores. The inhibitory •might be taken up and released by non- _ effect of LSD was abolished by phentol- adrenergic nerves, ,this is most unlikely in anaine (2 _g/ml) which itself markedly the case of [3H]-noradrenaline derived increased the contractions and efflux of from labelled tyrosine, i label during electrical stimulation The results with phentolamine may pro- i (Figure1). vide an explanationfor the potent and i Five experiments were carried out by prolonged effect of LSD. It has been Dr. Angela Waterfield on the isolated suggested that noradrenaline release may r" • longitudinal muscle-myenteric plexus be subject to a type of feed-back control preparation of the guinea-pig ileum. Con- exerted by noradrenaline itself on pre- tractions elicited by field stimulation at synaptic a-adrenoceptors (Farnebo & 0-1 Hz were rapidly depressed by LSD, a Mahnfors, 1971; Starke, 1972). LSD may 50% depression being obtained with mimic this effect by combining with the 100 ng/ml. Contractions of the myenteric presynaptic receptor sit_s. This would .... plexus elicited by acetylcholine were not explain why the effect of LSD is limited significantly affected by LSD. to very low frequencies or short trains of pulses, for under these conditions the intrinsic feed-back control would be less Discussion.---These results confirm and _ctive than when greater amounts of extend those of Ambache et al. (1971) and transmitter are released at higher frequen- Ambache, Dunk, Verney & Aboo Zar cies (Hughes, 1972) or longer trains of (1973): however, these workers consider pulses (Hughes & Roth, 1972). Noradrena- tha,t the motor innervation of the vas is line itself can depress the contractions at non-adrenergic. The present results sup- 0.1 Hz and this supports the concept of port the classical concept that noradrena- feed-back inhibition. The preliminary line is the motor transmitter in the vas results in the myenteric plexus provide deferens. The parallel reduction of both additional support for this hypothesis, for the contractions and label efflux during there is little doubt that acetylcholine is stimulation are most likely causally the transmitter here and it has been shown .. related. Although [3H]-noradrenaline that acetylcholine release is inhibited by 708 Short communications

a-adrenoceptor stimulation at this site BOAKES,R. J., BRADLEY,P. B., BR_GGS, I. & (Kosterlitz, Lydon & Watt, 11970). DRAY,A. (1970). Antagonism of 5-hydroxy- tryptamine by LSD-25 in the central It is possible that phentolamine may nervous system: a possible neuronal basis antagonize the effects of LSD not by pre- for the actions of LSD-25. Br. J. Pharmac., venting access to a presynaptic receptor 46, 202-218. site but by some other action, possibly FARNESO, L.-O. & MALMFORS, T. (1971). consequent upon the increased efflux of responseSH-noradrenalinein the fieldreleasestimulatedand mechanicalmouse vas transmitter. However, if LSD and phentol- deferens. Acta. physiol, scand., Suppl. amine do interact at the same receptor 371,1-18. _ then we may have to re-evaluate our con- GADDUM, J. H. (1957). Serotonin-LSD inter- t cept of _-adrenoceptors. LSD has c;nly a actions. Ann. N.Y. Acad. Sci., 66, 643-648. : weak action on vascular a-adrenoceptors HOFFMAN, A. (1968). Psychotomimetic agents. (Hoffman, 1968) and did not cause con- In: Drogs affecting the central nervous tractions of the guinea-pig vas deferens; system. Medicinal Research Series, Volume 2, pp. 169-226. London, Edward Arnold _. the presynaptic adrenoceptor would then Ltd. appear to" differ from the classical a-adrenoceptor in terms of its agonist HU_HcontrollingES, J. (1972).the releaseEvaluationand inactivationof mechanismsof specificity, the adtenergictransmitter in the rabbit REFERENCES portal vein and vas deferens. Br. J. Pharmac., 44, 472-491. i AMBACHE,N., DUNK, L. P., MIALL, P. & AaOO ZAR, M. (1971). Unexplained inhibitory HUGHES,J. & ROTH, R. H. (1972). Variations _ action of D-lysergic acid diethylamide in noradrenaline output with respect to (LSD) on postganglionic motor tram- stimulus frequency, train length and origin mission in the guinea-pig vas deferens, of the transmitter. Br. J. Pharmac., 45, Br. J. Pharmac., 42, 659-660P. 157P. AMBACHE, N., DUNK, L. P., VERNEY, J. & KOSTERLITZ, H. W., LYDON, R. J. & WATr, ABoo ZAR, M. (1973). An inhibition of A.J. (1970). The effects of adrenaline, post-ganglionic motor transmission in the noradrenaline and isoprenaline on inhibi- mammalian vas deferens by D-lysergic acid tory a- and fl-adrenoceptors in the longi- diethylamide. J. Physiol., Lond.; 231, tudinal muscle of the guinea-pig ileum. 251-270. Br. J. Pharmac., 39, 398--413. AND_N, N.-E., CORRODI,H., FUXE, K. & HOK- STARKE,K. (1972). Influence of extracellular FELT, T. (1968). Evidence for a centrat noradrenaline on the stimulation-evoked 5-hydroxytryptamine receptor stimulation secretion of noradrenaline from sympa- by lysergic acid diethylallnide. Br. J. thetic nerves: evidence for an a-receptor- Pharmac., 34, 1-7. mediated feedback inhibition of nor- IBOADLE-BIBER, M. C., HUGHES, J. & ROTH, adrenaline release. Natmyn-Schmiedeberg's R. H. (1970). Acceleration of noradrenaline Arch. Pharmac., 2,75, 11-23. biosynthesis in the guinea-pig vas deferens "' by potassium. Br. J. Pharmac., 40, 702-720. (Received June 26,1973)