The suprachiasmatic nucleus (SCN) is the principal circadian pacemaker in mammals. A salient feature of the SCN is that cells of a particular phenotype are topographically organized; this organization defines functionally distinct subregions that interact to generate coherent rhythmicity. In Syrian hamsters (Mesocricetus auratus), a dense population of directly retinorecipient calbindin D(28K) (CalB) neurons in the caudal SCN marks a subregion critical for circadian rhythmicity. In mouse SCN, a dense cluster of CalB neurons occurs during early postnatal development, but in the adult CalB neurons are dispersed through the SCN. In the adult retina CalB colocalizes with melanopsin-expressing ganglion cells. In the present study, we explored the role of CalB in modulating circadian function and photic entrainment by investigating mice with a targeted mutation of the CalB gene (CalB-/- mice). In constant darkness (DD), CalB-/- animals either become arrhythmic (40%) or exhibit low-amplitude locomotor rhythms with marked activity during subjective day (60%). Rhythmic clock gene expression is blunted in these latter animals. Importantly, CalB-/- mice exhibit anomalies in entrainment revealed following transfer from a light : dark cycle to DD. Paradoxically, responses to acute light pulses measured by behavioral phase shifts, SCN FOS protein and Period1 mRNA expression are normal. Together, the developmental pattern of CalB expression in mouse SCN, the presence of CalB in photoresponsive ganglion cells and the abnormalities seen in CalB-/- mice suggest an important role for CalB in mouse circadian function.