The suprachiasmatic nucleus (SCN), the central circadian clock in mammals, is a neural network consisting of various types of GABAergic neurons, which can be differentiated by the co-expression of specific peptides such as vasoactive intestinal peptide (VIP) and arginine vasopressin (AVP). VIP has been considered as a critical factor for the circadian rhythmicity and synchronization of individual SCN neurons. However, the precise mechanisms of how VIP neurons regulate SCN circuits remain incompletely understood. Here, we generated ViptTA knock-in mice that express tetracycline transactivator (tTA) specifically in VIP neurons by inserting tTA sequence at the start codon of Vip gene. The specific and efficient expression of tTA in VIP neurons was verified using EGFP reporter mice. In addition, combined with Avp-Cre mice, ViptTA mice enabled us to simultaneously apply different genetic manipulations to VIP and AVP neurons in the SCN. Immunostaining showed that VIP is expressed at a slightly reduced level in heterozygous ViptTA mice but is completely absent in homozygous mice. Consistently, homozygous ViptTA mice showed impaired circadian behavioral rhythms similar to those of Vip knockout mice, such as attenuated rhythmicity and shortened circadian period. In contrast, heterozygous mice demonstrated normal circadian behavioral rhythms comparable to wild-type mice. These data suggest that ViptTA mice are a valuable genetic tool to express exogenous genes specifically in VIP neurons in both normal and VIP-deficient mice, facilitating the study of VIP neuronal roles in the SCN neural network.

The central circadian clock of the suprachiasmatic nucleus (SCN) is a network consisting of various neurons and glia. Individual cells have the autonomous molecular machinery of a cellular clock, but their intrinsic periods are considerably variable. Here, we show that arginine vasopressin (AVP) neurons set the ensemble period of the SCN network to control circadian behavior rhythm. Artificial lengthening of cellular periods by deleting casein kinase 1 delta (CK1δ) in the whole SCN lengthened the free-running period of behavior rhythm to an extent similar to CK1δ deletion specific to AVP neurons. In SCN slices, PER2::LUC reporter rhythms of these mice did not recapitulate the period lengthening. However, in vivo calcium rhythms of both AVP and vasoactive intestinal peptide (VIP) neurons demonstrated lengthened periods similar to the behavioral rhythm upon AVP neuron-specific CK1δ deletion. These results indicate that AVP neurons act as the primary determinant of the SCN ensemble period.