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Intraocular injection of the Bartha strain of pseudorabies virus (PRV Bartha) results in transsynaptic infection of the hypothalamic suprachiasmatic nucleus (SCN), a retinorecipient circadian oscillator. PRV Bartha infection of a limited number of retinorecipient structures, including the SCN, was initially interpreted as the differential infection of a subpopulation of rat retinal ganglion cells, followed by replication and anterograde transport via the optic nerve. A recent report that used a recombinant strain of PRV Bartha (PRV152) expressing enhanced green fluorescent protein demonstrated that SCN infection actually results from retrograde transneuronal transport of the virus via the autonomic innervation of the eye in the golden hamster. In the present study using the rat, the pattern of infection after intravitreal inoculation with PRV152 was examined to determine if infection of the rat SCN is also restricted to retrograde transsynaptic transport. It was observed that infection in preganglionic autonomic nuclei (i.e., Edinger-Westphal nucleus, superior salivatory nucleus, and intermediolateral nucleus) precedes infection in the SCN. Sympathetic superior cervical ganglionectomy did not abolish label in the SCN after intraocular infection, nor did lesions of parasympathetic preganglionic neurons in the Edinger-Westphal nucleus. However, combined Edinger-Westphal nucleus ablation and superior cervical ganglionectomy eliminated infection of the SCN. This observation allowed a detailed examination of the SCN contribution to descending autonomic circuits afferent to the eye. The results indicate that in the rat, as in the hamster, SCN infection after intraocular PRV152 inoculation is by retrograde transsynaptic transport via autonomic pathways to the eye. J. Comp. Neurol. 471:298–313, 2004. © 2004 Wiley-Liss, Inc.

Melanopsin is a novel opsin synthesized in a small subset of retinal ganglion cells. Ganglion cells expressing melanopsin are capable of depolarizing in response to light in the absence of rod or cone input and are thus intrinsically light sensitive. Melanopsin ganglion cells convey information regarding general levels of environmental illumination to the suprachiasmatic nucleus, the intergeniculate leaflet, and the pretectum. Typically, retinal ganglion cells communicate information to central visual structures by receiving input from retinal photoreceptors via bipolar and amacrine cells. Because melanopsin ganglion cells do not require synaptic input to generate light-induced signals, these cells need not receive synapses from other neurons in the retina. In this study, we examined the ultrastructure of melanopsin ganglion cells in the mouse retina to determine the type (if any) of synaptic input these cells receive. Melanopsin immunoreaction product was associated primarily with the plasma membrane of (1) perikarya in the ganglion cell layer, (2) dendritic processes in the inner plexiform layer (IPL), and (3) axons in the optic fiber layer. Melanopsin-immunoreactive dendrites in the inner (ON) region of the IPL were postsynaptic to bipolar and amacrine terminals, whereas melanopsin dendrites stratifying in the outer (OFF) region of the IPL received only amacrine terminals. These observations suggested that rod and/or cone signals may be capable of modifying the intrinsic light response in melanopsin-expressing retinal ganglion cells.

The suprachiasmatic nucleus (SCN), a circadian oscillator, receives glutamatergic afferents from the retina and serotonergic (5-HT) afferents from the median raphe. 5-HT1B and 5-HT7 receptor agonists inhibit the effects of light on SCN circadian activity. Electron microscopic (EM) immunocytochemical procedures were used to determine the subcellular localization of 5-HT1B and 5-HT7 receptors in the SCN. 5-HT1B receptor immunostaining was associated with the plasma membrane of thin unmyelinated axons, preterminal axons, and terminals of optic and nonoptic origin. 5-HT1B receptor immunostaining in terminals was almost never observed at the synaptic active zone. To a much lesser extent, 5-HT1B immunoreaction product was noted in dendrites and somata of SCN neurons. 5-HT7 receptor immunoreactivity in γ-aminobutyric acid (GABA), vasoactive intestinal polypeptide (VIP), and vasopressin (VP) neuronal elements in the SCN was examined by using double-label procedures. 5-HT7 receptor immunoreaction product was often observed in GABA-, VIP-, and VP-immunoreactive dendrites as postsynaptic receptors and in axonal terminals as presynaptic receptors. 5-HT7 receptor immunoreactivity in terminals and dendrites was often associated with the plasma membrane but very seldom at the active zone. In GABA-, VIP-, and VP-immunoreactive perikarya, 5-HT7 receptor immunoreaction product was distributed throughout the cytoplasm often in association with the endoplasmic reticulum and the Golgi complex. The distribution of 5-HT1B receptors in presynaptic afferent terminals and postsynaptic SCN processes, as well as the distribution of 5-HT7 receptors in both pre- and postsynaptic GABA, VIP, and VP SCN processes, suggests that serotonin plays a significant role in the regulation of circadian rhythms by modulating SCN synaptic activity. J. Comp. Neurol. 432:371–388, 2001. © 2001 Wiley-Liss, Inc.

The afferent connections of the hypothalamic suprachiasmatic nucleus (SCN) of the golden hamster were examined using horseradish peroxidase (HRP) as the retrograde tracer molecule. Unilateral iontophoretic deposition of HRP into the SCN labeled ganglion cells bilaterally in the retinae. The labeled ganglion cells all had large somata and were randomly distributed across the retina. A similar number were labeled in each retina, which contrasted with the findings from injections into the optic chiasm and lateral geniculate body. Chiasm and geniculate injections both labeled three classes of ganglion cell (small, large, and giant) predominantly in the contralateral retinae. Telencephalic afferent projections to the SCN included the ventral subicular cortex and the septum. Notable diencephalic afferents included the dorsal lamina of the internal division of the ventral lateral geniculate nucleus (vLGN); the ipsilateral input was twice that of the contralateral projection. The same region of the vLGN was also noted to be reciprocally connected to the contralateral vLGN. The thalamic paraventricular nucleus was also heavily labeled but only ipsilaterally. Of functional significance, the SCN was discovered to innervate its contralateral homologue. Other less numerous afferents in the midbrain included the dorsal and median raphe nuclei and the dorsal nucleus of the lateral lemniscus. The afferent projections to the SCN determined in this study are discussed in regard to the known physiological role of the SCN as part of the circadian clock system.

The immunohistochemical localization of luteinizing hormone-releasing hormone (LHRH) was studied in paraffin and vibratome-sectioned tissue from adult female hamsters that were treated neonatally with monosodium glutamate (MSG) or hypertonic saline. There appeared to be a reduction in LHRH-positive fibers in the median eminence of animals with an MSG-induced lesion of the arcuate nucleus in paraffin-embedded tissue. However, when unembedded tissue was cut on a vibratome, the distribution of LHRH-positive fibers and perikarya was similar in both groups of animals. Fibers were seen coursing through the periventricular area and lateral hypo-thalamus to the median eminence. In addition, LHRH-positive fibers were seen in the organum vasculosum of the lamina terminalis, subfornical organ, septal and preoptic areas, fasciculus retroflexus, habenular complex, and several regions in the basal forebrain. Animals that were pretreated with colchicine had LHRH-positive perikarya in the medial habenular nucleus, diagonal band of Broca, and the medial olfactory tract.