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mise à jour du
25 octobre 2009
Pharmacol Biochem Behav
The new muscarinic M1-receptor agonist YM796 evokes yawning
and increases oxytocin secretion
from the posterior pituitary gland in rats
Fujikawa M, Yamada K, Nagashima M, Domae M, Furukawa T
Department of Pharmacology, Fukuoka University, Japan


The present experiments were performed to examine the effects of a new muscarinic M1-receptor agonist, (-)-YM796 ((-)-S-2,8-dimethyl-3-methylene-1-oxa-8-azaspiro[4.5]decane L-tartrate monohydrate), on yawning and oxytocin secretion from the posterior pituitary gland in rats YM796, at doses of 2.5-50 mg/kg (SC), elicited yawning.
The yawning response was markedly increased by pretreatment with a beta-adrenoceptor antagonist, pindolol (20 mg/kg, IP), which per se did not elicit yawning. The yawning induced by YM796 (10 mg/kg, SC) in combination with pindolol (20 mg/kg, IP) was inhibited by scopolamine (0.5 mg/kg, SC), a muscarinic receptor antagonist, and pirenzepine (300 micrograms/ rat, ICV) and EEDQ (N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline) (5 mg/kg, IP), muscarinic M1-receptor antagonists, but not by spiperone (0.5 mg/kg, SC), a dopamine D2-receptor antagonist, 4-DAMP (4-diphenylacetoxy-N-methylpiperidine methiodide) (100 micrograms/rat, ICV), a muscarinic M3-receptor antagonist, and [d(CH2)5, Tyr(Mc)2, Orn8]-vasotocin (100 ng/rat, ICV), an oxytocin receptor antagonist. YM796 at 2.5-50 mg/kg (SC) did not exert an action on prolactin levels but increased oxytocin secretion from the posterior pituitary gland in rats.
This augmentation of oxytocin secretion by YM796 was inhibited by scopolamine (0.5 mg/kg, SC) and pirenzepine (3 mg/kg, SC), but not by mecamylamine (1 mg/kg, IP), a nicotinic receptor antagonist. The present findings obtained with YM796 suggest that the muscarinic M2-receptor stimulation participates in causing yawning behavior and oxytocin secretion in rat.

PREVIOUS behavioral studies, including our experimental findings, have shown that the yawning behavior was induced by dopamine D2-receptor agonists and was blocked by both dopamine D,-receptor and muscarinic receptor antagonists, thereby indicating that the behavior involves dopaminergic and cholinergic activation. Cholinesterase inhibitors and muscarinic receptor agonists also induce yawning behavior, which is blocked by muscarinic receptor antagonists. but not by dopamine receptor antagonists, suggesting that the dopaminergic activation precedes cholinergic one. Thus, the dopaminergic-cholinergic activation seems to participate essentially in eliciting yawning behavior. Besides, since administration of oxytocin has been reported to cause yawning, oxytocinergic neuronal activities seems to be participated in producing the yawning behavior. On the other hand, intraventricular administration of bethanechol or carbachol produces the sustained release of oxytocin, which is abolished by atropine, but not by mecamyl-amine or hexamethonium. In cultured bovine granulosa, continuous exposure to acetylcholine resulted in a dosedependent increase in oxytocin secretion, which was completely abolished by the specific muscarinic receptor antagonists, atropine and scopolamine, but not by nicotinic receptor antagonists, implying that cholinergic neurotransmitters play a direct role in the regulation of oxytocin release through muscarinic receptors.
The muscarinic receptors have been designated as M,- or M,-receptors, depending on whether they have high or low affinity for pirenzepine. Recently. molecular cloning studies have revealed the existence of five distinct muscarinic receptor proteins (M,-M,). but their functions are as yet tentative.
(-t)-YM796 was reported to exert muscarinic Ml-receptor agonistic activity in the central nervous system, with relatively weak muscarinic M,-receptor and/or muscarinic M,-receptor agonistic activity for behavioral changes at a high dose. Moreover, the (-)-isomer of YM796 having higher affinity for muscarinic M,-receptors than (-t)-YM796 was identified from the ratio of the IC,,, value.
To determine the possible involvement of the M, subtype of muscarinic receptor in yawning and oxytocin release, we examined whether a new muscarinic M,-receptor agonist, (-)- YM796, elicits yawning responses and affects oxytocin
(-)-YM796, a novel muscarinic M,-receptor agonist, was capable of inducing yawning in this study. Thus, the present result confirms the previous reports describing that RS-86 and AFlO2B, putative muscarinic M,-receptor agonists. induce yawning. As for the potency of producing yawning, the frequencies were 6.8 -t 2.2 yawns in 60 min by YM796, a muscarinic M,-receptor agonist, alone at 10 mgikg SC and 20.4 -C 3.0 yawns by YM796 in combination with pindolol in this study. The frequency produced by pilocarpine, a muscarinic M,- and M,-receptor agonist. was 3.1 + 0.6 yawns in 60 min at 4 mg/kg, IP alone and 20.9 + 3. I yawns in combination with pindolol under similar experimental conditions in our previous reports (28). Studies on the agents that act selectively on muscarinic M,-receptors have not been reported. Accordingly, the muscarinic M,-receptors are definitely involved in causing yawning, but the possible involvement of muscarinic MZ- and M,-receptors may not be dismissed at present.
In our previous reports blockade of B-adrenoceptors caused by central B-adrenoceptor antagonists, pindolol. and others, which reach the brain through the blood-brain barrier, but not by peripheral B-adrenoceptor antagonists, facilitates the occurrence of yawning induced by dopaminergic agonists. such as apomorphine and talipexole (B-HT 920). Administration of LY-78335 and IJK-I 187A, phenylethanolamine-Nmethyltransferase inhibitors, which decrease adrenaline formation without affecting dopamine and noradrenaline levels in the brain, similarly potentiates the incidence of yawning as does B-adrenoceptor blockade. Therefore. the central adrenergic neuronal system was proposed to participate in the regulation of yawning for suppressive direction via padrenoceptors.
The yawning induced by YM796 was also potentiated by treatment with the P-adrenoceptor antagonist, pindolol, in the present study. The yawning induced by muscarinic receptor agonists, AF102B and RS-86, has been reported to be blocked by muscarinic receptor antagonists but to be unaffected by dopamine receptor antagonists. In the present study, the yawning behavior elicited by YM796 administered after pindolol was inhibited by scopolamine. a muscarinic receptor antagonist, and was also antagonized by pirenzepine and EEDQ, muscarinic M,-receptor antagonists, without being affected by 4-DAMP, a muscarinic M?-receptor antagonist. and spiperone. a dopamine D,-receptor antagonist. Thus, muscarinic M,-receptors are involved in the induction of the yawning behavior. Nicotinic receptor stimulation causes an increase of oxytotin release, and the muscarinic pathway also seems to be involved in regulating oxytocin release. In the present study. serum oxytocin levels were increased in a dose-dependent manner not only by nicotine but also by YM796. indicating the participation of both nicotinic and muscarinic receptor mechanisms.
The elevation of oxytocin levels produced by YM796 was inhibited by scopolamine, a muscarinic receptor antagonist. but not by mecamylamine. a nicotinic receptor antagonist. thus implying that the muscarinic receptor stimulation participates in the acceleration of oxytocin release from the posterior pituitary gland. In further experiments. pirenzepine, a muscarinic M,-receptor antagonist. did not affect the static oxytocin levels but antagonized the elevation of the oxytocin level by YM796. These findings indicate that muscarinic M,-receptors play a role in increasing oxytocin release. As for possible involvement of oxytocinergic neuronal activities in the yawning, central administration of oxytocin has been reported to elicit yawning behavior. Therefore, it might be a problem whether or not the release of oxytocin found in the plasma is related to the yawning response. In this study, the yawning induced by YM796 after pindolol was not prevented by the oxytocin receptor blockade. Accordingly, the release of oxytocin from the neurohypophysis after YM796 may not be related to the yawning response.
This would also suggest that central and peripheral oxytocin release is differentially regulated and YM796 acts at another level in the central nervous structures involved in the regulation and/or expression of this behavior. Moreover, as described in the introduction, the dopaminergic-cholinergic neuronal linkage seems to be essentially involved in eliciting yawning behavior. In fact, YM796-induced yawning was inhibited by muscarinic antagonists, but not by a dopamine D,-receptor antagonist in the present study. There has been also the proposal that the expression of yawning induced by dopaminergic agonists involves dopamine-oxytocin, but not oxytocin-dopamine, neuronal linkage. Besides, the oxytocin-elicited yawning behavior is inhibited by oxytocin receptor antagonists and muscarinic receptor antagonists, but not by dopamine receptor antagonists. In this study, the YM796-induced yawning was antagonized by muscarinic M,-receptor antagonists, but not by the oxytocin receptor antagonist.
Therefore, the findings suggest that the oxytocinergic mechanism precedes the muscarinic one and is in position between dopaminergic and muscarinic mechanism in producing yawning behavior. The present findings suggest that the muscarinic M,-receptor stimulation is involved in the occurrence of yawning behavior and secretion of oxytocin from the posterior pituitary gland in rats.