CIRCADIAN RHYTHM OF NEURONAL ACTIVITY IN SUPRACHIASMATIC NUCLEUS SLICES FROM THE VASOPRESSIN-DEFICIENT BRATTLEBORO RAT C. D. INGRAM,*† R. K. SNOWBALL* and R. MIHAI‡ *Neuroendocrine Research Group, Department of Anatomy, University of Bristol, Bristol BS8 1TD, U.K. ‡Department of Endocrinology, ‘‘Carol Davila’’ University of Medicine and Pharmacy, Bucharest, 71279, Romania Abstract––In vitro extracellular recordings were made from tissue slices of suprachiasmatic nucleus from homozygous Brattleboro rats which are deficient in vasopressin. A high proportion (56%) of neurons were excited by application of exogenous vasopressin, indicating that the V 1 receptors expressed by these neurons were functional. Basal activity of these vasopressin-sensitive neurons showed a marked circadian variation (higher during the subjective light phase) while vasopressin-insensitive neurons showed no significant variation, suggesting the presence of the V 1 receptor identifies a population of highly circadian neurons. Suprachiasmatic neurons from both homozygous rats and their heterozygous (vasopressin- containing) litter mates displayed a circadian rhythm of spontaneous (basal) activity, with firing rates declining during the subjective dark phase, indicating that the endogenous pacemaker driving the circadian rhythm was not dependent upon the presence of vasopressin. However, the peak of spontaneous activity displayed during the subjective light phase was significantly lower in the vasopressin-deficient animals. These data show that the presence of endogenous vasopressin within the suprachiasmatic nucleus is not necessary for the generation of the circadian pattern of activity. However, vasopressin does function to amplify the rhythm by its excitatory effect during the light phase. Copyright  1996 IBRO. Published by Elsevier Science Ltd. Key words: vasopressin, vasotocin, Brattleboro, suprachiasmatic nucleus, circadian rhythm, neurohypo- physeal peptides. Neurons of the hypothalamic suprachiasmatic nu- clei (SCN) display a circadian cycle of electrical activity which peaks during the subjective light phase. The mechanisms generating this cycle appear to be intrinsic to the nucleus, since it persists after isolation in vitro. 10,11,18,25,27 However, the neuro- chemical signals regulating this rhythm are poorly understood. 13,31 Neurons in the dorsomedial area of the SCN synthesize and secrete [Arg 8 ]vasopressin (AVP), which is the only endogenous transmitter so far demonstrated to display a persistent, intrinsic rhythm of expression and release in vitro. 4,8,10,20 This release of AVP appears to play an important role in the output of the SCN, and has been suggested to be involved in determining circadian patterns of corti- costerone secretion, 14 and motor 1,3,15 and appeti- tive 22,23 behaviours. In addition to being involved as part of the output from the SCN, the remarkable correlation between the in vitro cycles of AVP release 8,10 and of electrical activity 10,11,25,27 (both peaking in the middle of the light phase) has led to the hypothesis that the endogenous cycle of AVP release contributes to the cycle of electrical activity and, thereby, functions as an integral part of the circadian clock mechanism. This is supported by the demonstration that application of an AVP receptor antagonist reduces spontaneous activity, 18 with a greater effect during the light phase, 19 indicating the presence of an endogenous AVP excitation which alters with the phase of the cycle. However, whether this endogenous excitatory effect can entirely account for the circadian changes in electrical activity requires that the cycle of activity be measured in the absence of endogenous AVP. The Brattleboro strain of Long–Evans rats carries a single base deletion in Exon B of the AVP gene. 24 In the homozygous animal, this recessive mutation results in an inability to process the translated gene product to AVP. Immunocytochemical staining has shown the complete absence of AVP-like material in the SCN of these animals. 32 Nevertheless, in spite of this deficiency, homozygous Brattleboro rats con- tinue to display circadian rhythms of cortical electro- encephalogram, 2 motor activity, 12,21 drinking behaviour 21,29 and pineal N-acetyltransferase lev- els. 21 These observations have led to the conclusion †To whom correspondence should be addressed. Abbreviations: AVP, [Arg 8 ]-vasopressin; AVT, [Arg 8 ]- vasotocin; GMT, Greenwich Mean Time; SCN, suprachi- asmatic nucleus; ZT, zeitgeber time. Pergamon Neuroscience Vol. 75, No. 2, pp. 635–641, 1996 Copyright  1996 IBRO. Published by Elsevier Science Ltd Printed in Great Britain 0306–4522/96 $15.00+0.00 PII: S0306-4522(96)00274-6 635