Circadian clocks are widely distributed in mammalian tissues, but little is known about the physiological functions of clocks outside the suprachiasmatic nucleus of the brain. The retina has an intrinsic circadian clock, but its importance for vision is unknown. Here we show that mice lacking Bmal1, a gene required for clock function, had abnormal retinal transcriptional responses to light and defective inner retinal electrical responses to light, but normal photoreceptor responses to light and retinas that appeared structurally normal by light and electron microscopy. We generated mice with a retina-specific genetic deletion of Bmal1, and they had defects of retinal visual physiology essentially identical to those of mice lacking Bmal1 in all tissues and lacked a circadian rhythm of inner retinal electrical responses to light. Our findings indicate that the intrinsic circadian clock of the retina regulates retinal visual processing in vivo.

The synaptic connections of the narrow-field, bistratified rod amacrine cell (AII) in the inner plexiform layer (IPL) of the rabbit retina were reconstructed from electron micrographs of continuous series of thin sections. The AII amacrine cell receives a large synaptic input from the axonal endings of rod bipolar cells in the most vitreal region of the IPL (sublamina b, S5) and a smaller input from axonal endings of cone bipolar cells in the scleral region of the IPL (sublamina a, S1-S2). Amacrine input, localized at multiple levels in the IPL, equals the total number of synapses received from bipolar cells. The axonal endings of cone bipolar cells represent the major target for the chemical output of the AII amacrine cell: these synapses are established by the lobular appendages in sublamina a (S1-S2). Ganglion cell dendrites represent only 4% of the output of the AII amacrine and most of them are also postsynaptic to the cone bipolars which receive AII input. The AII amacrine is not presynaptic to other amacrine cells. Finally, the AII amacrine makes gap junctions with the axonal arborizations of cone bipolars that stratify in sublamina b (S3-S4) as well as with other AII amacrine cells in S5. Therefore, in the rabbit retina 1) the rod pathway consists of five neurons arranged in series: rod-->rod bipolar-->AII amacrine-->cone bipolar-->ganglion cell; 2) it seems unlikely that a class of ganglion cells exists that is exclusively devoted to scotopic functions. In ventral, midperipheral retina, about nine rod bipolar cells converge onto a single AII amacrine, but one of them establishes a much higher proportion of synaptic contacts than the rest. Conversely, each rod bipolar cell diverges onto four AII amacrine cells, but one of them receives the largest fraction of synapses. Thus, within the pattern of convergence and divergence suggested by population studies, preferential synaptic pathways are established.

Anatomical and electrophysiological techniques were combined to study the morphology, synaptic connections, and response properties of two neurons in the rod pathway of the rabbit retina: the rod bipolar cell and the narrow-field, bistratified (NFB) amacrine cell. Rod bipolars receive synaptic input from rod cells in the outer plexiform layer (OPL), where their dendrites end as central elements in the invaginating synapse of rod spherules. Their main synaptic output in the inner plexiform layer (IPL) is onto NFB amacrine cells and at least one other type of amacrine, which in turn feeds a reciprocal synapse back onto the bipolar endings. Rod bipolars, or a variety of them, respond to diffuse, white light stimulation with a transient-sustained depolarization dominated by rods; with high-intensity flashes, they generate a secondary depolarization at off, which is homologous to the rod aftereffect of horizontal cells, although opposite in polarity. NFB amacrine cells receive synaptic input from rod bipolars, cone bipolars, and other types of amacrine cells; they are presynaptic to ganglion cell dendrites and communicate via gap junctions with other processes, whose parent neuron has not yet been identified. They respond to light with a triphasic potential, characterized by a depolarizing transient at on, followed by a sustained plateau phase, and finally by a hyperpolarizing transient at off. Threshold of their responses is the same as in the depolarizing rod bipolars and saturation is reached with nearly the same stimulus intensity in both neurons. Furthermore, NFB amacrine cells exhibit a depolarizing rod aftereffect at the termination of high-intensity flashes. Thus, this amacrine cell type is inserted in series along the rod pathway in the rabbit retina and modulates the transfer of scotopic signals from rod bipolars to ganglion cells.

We used a preparation of acutely dissociated neurons to quantify the ionic currents driving the spontaneous firing of substantia nigra pars compacta neurons, isolated from transgenic mice in which the tyrosine hydroxylase promoter drives expression of human placental alkaline phosphatase (PLAP) on the outer surface of the cell membrane. Dissociated neurons identified by fluorescent antibodies to PLAP showed firing properties similar to those of dopaminergic neurons in brain slice, including rhythmic spontaneous firing of broad action potentials and, in some cells, rhythmic oscillatory activity in the presence of tetrodotoxin (TTX). Spontaneous activity in TTX had broader, smaller spikes than normal pacemaking and was stopped by removal of external calcium. Normal pacemaking was also consistently silenced by replacement of external calcium by cobalt and was slowed by more specific calcium channel blockers. Nimodipine produced a slowing of pacemaking frequency. Pacemaking was also slowed by the P/Q-channel blocker -Aga-IVA, but the N-type channel blocker -conotoxin GVIA had no effect. In voltage-clamp experiments, using records of pacemaking as command voltage, cobalt-sensitive current and TTX-sensitive current were both sizeable at subthreshold voltages between spikes. Cobalt-sensitive current was consistently larger than TTX-sensitive current at interspike voltages from Ϫ70 to Ϫ50 mV, with TTX-sensitive current larger at voltages positive to Ϫ45 mV. These results support previous evidence for a major role of voltage-dependent calcium channels in driving pacemaking of midbrain dopamine neurons and suggest that multiple calcium channel types contribute to this function. The results also show a significant contribution of subthreshold TTX-sensitive sodium current.

The mesencephalic dopaminergic (mDA) cell system is composed of two major groups of projecting cells in the substantia nigra