NICOTINE PHASE-ADVANCES THE CIRCADIAN NEURONAL-ACTIVITY RHYTHM IN RATSUPRACHIASMATIC NUCLEI EXPLANTS

In vivo studies reported that cholinergic agents affect mammalian circ adian rhythmicity. To study phase resetting properties of cholinergic compounds more directly, we carried out experiments in rat suprachiasm atic nuclei slices. Compounds were added to the perfusate for Ih at sp ecific phases of the circadian cycle. On the following day, the time o f peak neuronal activity, a measure of the phase of the endogenous cir cadian pacemaker, was assessed by means of extracellular recording in the suprachiasmatic nuclei. The peak of neuronal activity occurred at circadian time 5.8 +/- 0.7 (mean +/- 95% confidence limits) in the con trol slice (circadian time 0: lights-on). Ten-micromolar carbachol had no effect on the phase of the circadian rhythm when given at circadia n times 6 and 15, while at circadian time 21 a phase advance of one ho ur was observed. By contrast, 10 mu M nicotine significantly phase adv anced (> 1 h) the neuronal circadian rhythm at all but one experimenta l circadian phase. The circadian times of maximal nicotinic phase adva nces were 15 (+2.6 h) and 21 (+2.8 h). A concentration response curve for nicotine was generated and pharmacological blocking experiments we re performed at circadian time 15. The estimated maximum response of n icotine was 3.4 h, and the estimated concentration for half maximal re sponse was 5 mu M. The Hill coefficient (=1.08) indicated that the eff ects of nicotine may be explained by a single receptor occupancy model . Mecamylamine (20 mu M) almost completely antagonized the nicotinic p hase-advances, whereas tetrodotoxin (1 mu M) or high Mg2+ (10 mM) did not significantly attenuate the nicotinic phase-advances. We conclude that nicotine given at most circadian phases advances the endogenous c ircadian clock. The efficacious blockade by mecamylamine, a classical nicotinic acetylcholine receptor antagonist, and tetrodotoxin Mg2+-ins ensitivity suggest that the phase shifts were selectively mediated by postsynaptic nicotinic receptors on putative clock cells in the suprac hiasmatic nuclei. The pharmacological properties of a recently charact erized alpha 7 subunit-bearing nicotinic receptor in the hypothalamus suggest that the present nicotinic phase advances in the suprachiasmat ic nuclei are mediated through binding at such receptors on putative c lock cells.