Serotonergic innervation of the hypothalamic suprachiasmatic nucleus and photic regulation of circadian rhythms

Converging lines of evidence have firmly established that the hypothalamic suprachiasmatic nucleus (SCN) is a light-entrainable circadian oscillator in mammals, critically important for the expression of behavioral and physiological circadian rhythms. Photic information essential for the daily phase resetting of the SCN circadian clock is conveyed directly to the SCN from retinal ganglion cells via the retinohypothalamic tract. The SCN also receives a dense serotonergic innervation arising from the mesencephalic raphe. The terminal fields of retinal and serotonergic afferents within the SCN are co-extensive, and serotonergic agonists can modify the response of the SCN circadian oscillator to light. However, the functional organization and subcellular localization of 5HT receptor subtypes in the SCN are just beginning to be clarified. This information is necessary to understand the role 5HT afferents play in modulating photic input to the SCN. In this paper, we review evidence suggesting that the serotonergic modulation of retinohypothalamic neurotransmission may be achieved via at least two different cellular mechanisms: 1) a postsynaptic mechanism mediated via 5HT1A or 5ht7 receptors located on SCN neurons; and 2) a presynaptic mechanism mediated via 5HT1B receptors located on retinal axon terminals in the SCN. Activation of either of these 5HT receptor mechanisms in the SCN by specific 5HT agonists inhibits the effects of light on circadian function. We hypothesize that 5HT modulation of photic input to the SCN may serve to set the gain of the SCN circadian system to light.     

Neuropeptide Y and enkephalin immunoreactivity in retinorecipient nuclei of the hamster pretectum and thalamus

This investigation was stimulated by the historical confusion concerning the identity of certain pretectal nuclei and by large differences reported between species with respect to which nuclei receive retinal innervation. Subcortical visual nuclei were studied using immunohistochemistry to identify retinal projections labeled following intraocular injection of cholera toxin, b fragment. In addition, neuropeptide Y (NPY) or enkephalin (ENK) immunoreactive cells and fibers were also evaluated in the retinorecipient pretectal and thalamic areas. The results confirm the established view that the retina directly innervates the nucleus of the optic tract (NOT), posterior (PPT), and olivary pretectal (OPT) nuclei. However, the retina also innervates the hamster medial (MPT) and anterior (APT; dorsal division) pretectal nuclei, results not previously reported in rodents. A commissural pretectal area (CPT) sparsely innervated by retina is also described. The data show for the first time that the posterior limitans nucleus (PLi) receives a moderately dense, direct retinal input. The PLi does not project to the cortex and appears to be a pretectal, rather than thalamic, nucleus. All retinal projections are bilateral, although predominantly contralateral. The PLi contains a moderately dense plexus of NPY- and ENK-IR fibers and terminals. However, peptidergic fibers also traverse the ATP and connect with the dorsomedial pretectium. The OPT contains ENK- and NPY-IR neurons and fibers, but is specifically identifiable by a moderately dense plexus of ENK-IR terminals. Numerous ENK-IR neurons are found in the NOT and PPT. The latter also has moderate numbers of ENK-IR fibers and terminals, but few NPY-IR neurons or fibers. The MPT contains modest numbers of ENK-IR fibers. The APT has no NPY-IR neurons or terminals, but an occasional ENK-IR neuron is seen and there is sparse ENK-IR innervation. Peptidergic innervation of the visual nuclei does not appear to be derived from the retina. The results show a set of retinally innervated, contiguous nuclei extending from the thalamic ventrolateral geniculate nucleus dorsomedially to the midbrain CPT. These nuclei plus the superior colliculus comprise a dorsal "visual shell" embracing a central core of caudal thalamus and rostral midbrain.     

Organization of neural inputs to the suprachiasmatic nucleus in the rat

The circadian timing of the suprachiasmatic nucleus (SCN) is modulated by its neural inputs. In the present study, we examine the organization of the neural inputs to the rat SCN using both retrograde and anterograde tracing methods. After Fluoro-Gold injections into the SCN, retrogradely labeled neurons are present in a number of brain areas, including the infralimbic cortex, the lateral septum, the medial preoptic area, the subfornical organ, the paraventricular thalamus, the subparaventricular zone, the ventromedial hypothalamic nucleus, the posterior hypothalamic area, the intergeniculate leaflet, the olivary pretectal nucleus, the ventral subiculum, and the median raphe nuclei. In the anterograde tracing experiments, we observe three patterns of afferent termination within the SCN that correspond to the photic/raphe, limbic/hypothalamic, and thalamic inputs. The median raphe projection to the SCN terminates densely within the ventral subdivision and sparsely within the dorsal subdivision. Similarly, areas that receive photic input, such as the retina, the intergeniculate leaflet, and the pretectal area, densely innervate the ventral SCN but provide only minor innervation of the dorsal SCN. A complementary pattern of axonal labeling, with labeled fibers concentrated in the dorsal SCN, is observed after anterograde tracer injections into the hypothalamus and into limbic areas, such as the ventral subiculum and infralimbic cortex. A third, less common pattern of labeling, exemplified by the paraventricular thalamic afferents, consists of diffuse axonal labeling throughout the SCN. Our results show that the SCN afferent connections are topographically organized. These hodological differences may reflect a functional heterogeneity within the SCN.     

Increased contact time improves adenovirus-mediated CFTR gene transfer to nasal epithelium of CF mice

By combining retrograde and anterograde tracing, evidence for a bineuronal connection from the suprachiasmatic nucleus (SCN) to the intermediolateral cell column in the spinal cord (IML) was obtained. The retrograde tracer cholera toxin subunit B (ChB) was pressure-injected into the spinal cord and the anterograde tracer Phaseolus vulgaris-leucoagglutinin (PHA-L) was iontophoretically injected into the SCN. The two tracers were visualized simultaneously by a double immunohistochemical procedure. In the hypothalamus, ChB injections gave rise to retrogradely labeled cell bodies in the paraventricular nucleus, retrochiasmatic area, perifornical region, lateral hypothalamic area, and the posterior hypothalamic area. The SCN were found to project to all of these areas. Furthermore, spinal-projecting neurons were found in the brain stem, but no efferents from the SCN were observed to innervate these areas. In the most sparsely innervated areas, the lateral hypothalamic area and the perifornical region, only occasionally a PHA-L fiber in close apposition to a ChB-ir cell body was observed. This was also the case in the retrochiasmatic area and posterior hypothalamic area, although these areas received a moderate number-immunoreactive (ir) PHA-L-ir fibers. The highest number of closely apposed PHA-L-ir fibers and ChB-ir cell bodies was observed in the dorsal parvicellular and in the ventral division of the medial parvicellular paraventricular nucleus, which were also the areas receiving the densest input from the SCN. By anterograde tracing from the paraventricular nucleus of the hypothalamus, the exact topography of the terminal field formed by descending paraventricular neurons was established. Thus, it was confirmed that the paraventricular nucleus of the hypothalamus predominantly innervates the IML. The present study suggests the existence of a bineuronal link between the SCN and the IML, possibly involved in transmission of circadian signals from the endogenous clock to the pineal gland and other organs receiving sympathetic afferents.     

Compliance in an anti-hypertension trial: a latent process model for binary longitudinal data

Animal experimental studies have shown that the retinohypothalamic tract (RHT) is an anatomical and functionally distinct retinofugal pathway mediating photic entrainment of circadian rhythms. In the present study, RHT projections were studied in the human brain by our recently developed postmortem tracing technique with neurobiotin as a tracer. Similar patterns of labeling were observed in brains of one control subject without neurological or mental disorders and five patients with Alzheimer's disease. The topography of RHT projections has several characteristics. (1) RHT fibers leave the optic chiasm and enter the hypothalamus medially and laterally at the anterior level of the suprachiasmatic nucleus (SCN). (2) The medial fibers enter the ventral part of the SCN and innervate the ventral SCN over its entire length, but the density decreases gradually from anterior to posterior. Labeled RHT fibers in the SCN make contact mainly with immunocytochemically positive neurotensin or vasoactive intestinal polypeptide neurons and only occasionally with vasopressin-positive neurons located in the ventral part of the SCN. (3) Only few projections to the dorsal part of the SCN and the anteroventral part of the hypothalamus were found. (4) Lateral projections reach the ventral part of the ventromedial SON and the area lateral to the SCN. No projections were observed to other hypothalamic areas. The presence of an RHT in humans suggests that the RHT may serve a function in humans similar to that demonstrated in animals.     

The suprachiasmatic nucleus and intergeniculate leaflet of Arvicanthis niloticus, a diurnal murid rodent from East Africa

Little is known about the neural substrates controlling circadian rhythms in day-active compared to night-active mammals primarily because of the lack of a suitable diurnal rodent with which to address the issue. The murid rodent, Arvicanthis niloticus, was recently shown to exhibit a predominantly diurnal pattern of activity and body temperature, and may be suitable for research on the neural mechanisms underlying circadian rhythms. This paper describes, in A. niloticus, the anatomy of two neural structures that play important roles in the control of circadian rhythms, the suprachiasmatic nucleus (SCN) and the intergeniculate leaflet (IGL). Immunohistochemical techniques were used to examine the distribution of neuroactive peptides in the SCN and IGL, and retinal projections to these structures were traced with anterograde transport of the beta subunit of cholera toxin. In A. niloticus, distinct subdivisions of the SCN contained cell bodies with immunoreactive (IR) vasopressin, vasoactive intestinal polypeptide, gastrin-releasing peptide, and corticotropin-releasing factor. The SCN did not contain cell bodies with met-enkephalin-IR and substance P-IR, but did contain fibers with substance P-IR and neuropeptide Y-IR. Retinal fibers were present throughout the SCN, but were most densely concentrated along its ventral edge, particularly in the contralateral SCN. Retinal fibers also extended to a variety of hypothalamic regions outside the SCN, including the supraoptic nucleus and the subparaventricular region. The IGL contained cells with neuropeptide Y-IR and enkephalin-IR cells. Retinal fibers projected to both the ipsilateral and contralateral IGL. The anatomy of the SCN and IGL were compared and contrasted with that previously described for other nocturnal and diurnal species.     

Photic entrainment of circadian rhythms in rodents

Photic entrainment of circadian rhythms occurs as a consequence of daily, light-induced adjustments in the phase and period of the suprachiasmatic nuclei (SCN) circadian clock. Photic information is acquired by a unique population of retinal photoreceptors, processed by a distinct subset of retinal ganglion cells, and conveyed to the SCN through the retinohypothalamic tract (RHT). RHT neurotransmission is mediated by the release of the excitatory amino acid glutamate and appears to require the activation of both NMDA- and non-NMDA-type glutamate receptors, the expression of immediate early genes (IEGs), and the synthesis and release of nitric oxide. In addition, serotonin appears to regulate the response of the SCN circadian clock to light through postsynaptic 5-HT1A or 5-ht7 receptors, as well as presynaptic 5-HT1B heteroreceptors on RHT terminals.     

A direct retinal projection to the dorsal raphe nucleus in the rat

A direct projection from the retina to the dorsal raphe nucleus at the pontomesencephalic junction was demonstrated with both antero- and retrograde tracing techniques in the rat. Following intravitreous injections of choleratoxin subunit B (CTB), horseradish peroxidase (HRP) and CTB-conjugated HRP, varicose fibers were labeled in the lateral region of the dorsal raphe nucleus, predominantly contralateral to the injection. Many of these labeled fibers were intermingled with serotonin-immunoreactive neurons, but some fibers were also found further laterally, beyond the boundary of dorsal raphe nucleus but within the periaqueductal gray. Following injections of the retrograde tracers Fluoro-Gold and CTB into the dorsal raphe nucleus and adjacent periaqueductal gray (without contamination of previously known targets of retinal projections), a small population of ganglion cells was labeled in the retina. These data provide evidence for the existence of a direct retinal projection to the lateral region of the dorsal raphe nucleus and the adjacent mesopontine periaqueductal gray in the rat. This projection may have a role in sensorimotor coordination and the regulation of circadian rhythm as well as sleep and wakefulness.     

Efferents from the suprachiasmatic nucleus to basal forebrain nuclei in the green treefrog (Hyla cinerea)

Injections of horseradish peroxidase (HRP) into the eye, striatum, preoptic area, or ventral hypothalamus of the green treefrog (Hyla cinerea) demonstrated an indirect retinal pathway to each basal forebrain region via the suprachiasmatic nucleus (SCN). Intraocular injections resulted in HRP-filled fibers within the ventral portion of the SCN bilaterally. Apparent en passant and terminal swellings on these fibers were seen adjacent to SCN cells. Cells in both the ventral and dorsal portions of the SCN were retrogradely filled following each of the forebrain injections, but the relative distributions of filled cells were different. The striatum receives a bilateral projection mainly from cells in the ventral portion of the SCN. A small ipsilateral projection from this same SCN region reaches the ventral hypothalamus. The SCN projection to the preoptic area arises from a more uniform distribution of cells, with the majority located in the ipsilateral dorsal region of the nucleus. The striatum is believed to be involved in orientation responses, while the preoptic area and ventral hypothalamus regulate gonadal activity related to reproduction. Each of these areas receives auditory input carrying information about acoustic communication signals, social cues important for coordinating reproductive activity. Input from the SCN could provide these same basal forebrain centers with information about the photic environment, an important environmental cue regulating reproduction.     

Neurons containing gastrin-releasing peptide and vasoactive intestinal polypeptide are involved in the reception of the photic signal in the suprachiasmatic nucleus of the Syrian hamster: an immunocytochemical ultrastructural study

In mammals, the suprachiasmatic nuclei are involved in the generation of biological rhythms and are synchronized by light input coming from the retina. The targets of retinal afferents and the involvement of neurons containing gastrin-releasing and vasoactive intestinal peptides in photic reception were investigated in the suprachiasmatic nuclei of the Syrian hamster by using light- and electron-microscopic immunocytochemistry. Cholera toxin was used to trace retinal fibers and Fos immunoreactivity to visualize cellular response to light stimulation. Ultrastructural observations were made in the intermediate third of the nuclei, the area of highest overlap for the immunoreactivities investigated. Gastrin-releasing peptide and vasoactive intestinal peptide cell bodies were localized in the ventral part of the nuclei; their dense immunoreactive fiber network often displayed synaptic contacts. Both neuropeptides were colocalized in elongated cells observed near the optic chiasm. Following a light pulse in the middle of the subjective night, Fos protein was expressed in most gastrin-releasing peptide perikarya and in some vasoactive intestinal peptide cells. Retinal terminals mostly occurred in the midline zone between the suprachiasmatic nuclei. Symmetrical or asymmetrical retinal synapses were observed on gastrin-releasing peptide-immunoreactive dendrites and somata, but never on vasoactive intestinal peptide neurons. These results are discussed in relation to the photic entrainment of the circadian clock.     

The suprachiasmatic nucleus: cellular approach to clock functioning

The hypothalamic suprachiasmatic nucleus (SCN), the circadian clock in mammals, generates and maintains a variety of daily rhythms. The present review is an attempt to synthesis experimental data on the anatomical organisation and cellular activities within SCN. The clock exhibits an endogenous rhythmic activity and can also be entrained by environmental synchronisers such as the light/dark cycle. It can be also influenced by internal signals such as the rhythmic secretion of melatonin which is under control of SCN activity. This tiny structure contains a variety of peptides organised in a specific distribution. It receives three main inputs from the retina (glutamate), the intergeniculate leaflet (NPY) and the dorsal raphe (serotonin). VIP containing cells located in the ventral part of SCN receive all these afferences and innervate the whole structure. VIP, PHI and GRP are likely implicated in the entrainment of the clock. The vasopressin (VP) cells exhibiting an endogenous rhythmic synthesis are considered as an output of the clock. The specific induction of immediate early genes (c-fos, jun B) within SCN by light pulses during the subjective night suggests the participation of these genes in the process of cellular entrainment by the photic input. The demonstration of a rhythmic astrocytic activity within SCN suggests an active involvement of this cellular population in the functioning of the clock facilitating or not neuronal communication. Cellular disturbances such as a decrease in VIP or VP cell population, reduction in the amplitude of functional cellular rhythms, astrocytic proliferation could explain some pathologies observed with ageing.     

Insular cortex and amygdala lesions induced after aversive training impair retention: effects of degree of training

Immunohistochemical observation was performed in the suprachiasmatic nucleus (SCN) and the intergeniculate leaflet (IGL) of hereditary bilaterally microphthalmic rats without the optic nerve on both sides. In the microphthalmic rats, volume of the SCN reduced to ca. 70% of the normal and numbers of the vasoactive intestinal polypeptide (VIP)-like immunoreactive (lir) neurons were significantly decreased. Although the arginine vasopressin (aVP)- and the VIP-lir neurons distributed in the dorsomedial and the ventrolateral part of the SCN, respectively, as reported in the normal one, somatostatin-lir neurons, localizing mainly in a border area between the dorsomedial and the ventrolateral region of the normal SCN, were shifted to the ventral part of the SCN in the microphthalmic rats. The ventral part of the SCN was covered with neuropeptide Y (NPY)-lir fibers in both normal and mutant rats. The IGL was hardly delineated cytologically in the lateral geniculate nucleus (LGN) of the mutant rats. NPY-lir neurons were found in the dorsal part of the ventral LGN, in contrast to their even distribution in the normal IGL. These findings suggest that the IGL-SCN tract remains in the hereditary microphthalmic rats without the retinal projections.     

Pattern of distribution of neuropeptides in the camel lacrimal gland

The pattern of distribution of neuropeptides, including neuropeptide-Y (NPY), vasoactive intestinal polypeptide (VIP), neurotensin (NT), serotonin (5-HT), galanin (GAL), leucine-enkephalin (LEU-ENK) and calcitoningene-related-peptide (CGRP), in the nerves of the camel lacrimal gland was investigated using immunohistochemical techniques. Fresh lacrimal gland segments, obtained from adult camels slaughtered in the local abattoir, were used for the immunohistochemical techniques. NPY and LEU-ENK immunoreactivity was observed in the nerve cell bodies and nerve fibers of the camel lacrimal gland. VIP, GAL and CGRP were demonstrated predominantly in fine varicose nerve fibers lying on the basolateral surfaces of the lacrimal acinar cells. NT and 5-HT were identified mainly in neurons situated in the periacinar regions, close to the basal surfaces of the acinar cells. It is concluded that the camel lacrimal nerves contain several neuropeptides including NPY, VIP, NT, 5-HT, GAL, LEU-ENK and CGRP which may modulate lacrimal fluid and protein secretion.     

Rapid resetting of the mammalian circadian clock

The suprachiasmatic nuclei (SCN) contain the principal circadian clock governing overt daily rhythms of physiology and behavior. The endogenous circadian cycle is entrained to the light/dark via direct glutamatergic retinal afferents to the SCN. To understand the molecular basis of entrainment, it is first necessary to define how rapidly the clock is reset by a light pulse. We used a two-pulse paradigm, in combination with cellular and behavioral analyses of SCN function, to explore the speed of resetting of the circadian oscillator in Syrian hamster and mouse. Analysis of c-fos induction and cAMP response element-binding protein phosphorylation in the retinorecipient SCN demonstrated that the SCN are able to resolve and respond to light pulses presented 1 or 2 hr apart. Analysis of the phase shifts of the circadian wheel-running activity rhythm of hamsters presented with single or double pulses demonstrated that resetting of the oscillator occurred within 2 hr. This was the case for both delaying and advancing phase shifts. Examination of delaying shifts in the mouse showed resetting within 2 hr and in addition showed that resetting is not completed within 1 hr of a light pulse. These results establish the temporal window within which to define the primary molecular mechanisms of circadian resetting in the mammal.     

GABAergic presubicular projections to the medial entorhinal cortex of the rat

We characterized presubicular neurons giving rise to bilateral projections to the medial entorhinal cortex (MEA) of the rat. Retrograde labeling of presubiculo-entorhinal projections with horseradish peroxidase and subsequent GABA immunocytochemistry revealed that 20-30% of the ipsilaterally projecting neurons are GABAergic. No GABAergic projections to the contralateral MEA were observed. GABAergic projection neurons were observed only in the dorsal part of the presubiculum, which, when taking into account the topography of presubicular projections to MEA, indicates that only the dorsal part of MEA receives GABAergic input. The GABAergic projection neurons constitute approximately 30-40% of all GABAergic neurons present in the superficial layers of the dorsal presubiculum. Using double-label fluorescent retrograde tracing, we found that the ipsilateral and contralateral presubiculo-entorhinal projections originate from different populations of neurons. Anterograde labeling of presubiculo-entorhinal projections and electron microscopical analysis of labeled terminals substantiated the presence of a restricted GABAergic presubiculo-entorhinal projection. A small fraction of afferents to only ipsilateral dorsal MEA formed symmetrical synapses with dendritic shafts. No symmetrical synapses on spines were noted. Most afferents to the dorsal part of ipsilateral MEA, as well as all afferents to the remaining ipsilateral and contralateral MEA, formed asymmetrical synapses with both spines and dendritic shafts in an almost equal ratio. Thus, we conclude that the majority of the presubiculo-entorhinal projections exert an excitatory effect on both principal neurons and interneurons. The projections from the dorsal part of the presubiculum comprise a small inhibitory component that originates from GABAergic neurons and targets entorhinal interneurons.     

Morphology, axonal projection pattern, and response types of tectal neurons in plethodontid salamanders. I: tracer study of projection neurons and their pathways

In three salamander species (Hydromantes italicus, H. genei, Plethodon jordani), the tectobulbospinal and tectothalamic pathways and their cells of origin were studied by means of anterograde and retrograde biocytin and tetramethylrhodamine tracing. In plethodontid salamanders, five types of tectal projection neurons were identified. TO1 neurons have widefield dendritic trees that arborize in the layers of retinal afferents and form a neuropil in the superficial layer; axons constitute the crossed tectospinal tract. Dendrites of TO2 cells have the largest dendritic trees that arborize in the intermediate and deep layers of retinal afferents; axons constitute a lateral uncrossed tectospinal tract. TO3 cells have widefield dendritic trees that arborize in the deep layer of retinal afferents and in the layer of tectal efferents; axons constitute a superficial uncrossed tectospinal tract. TO4 cells have slender primary dendrites and small-field dendritic trees that arborize in the intermediate layers of retinal afferents; axons constitute another lateral uncrossed tectospinal tract. TO2, TO3, and TO4 cells also have ascending axons that run to the ventral and dorsal thalamus. TO5 cells have slender primary dendrites and small-field dendritic trees that extend into the superficial layers of retinal afferents; their fine axons constitute the bulk of the pathways ascending to the ipsilateral and contralateral thalamus. These morphological types of projection neurons and their ascending and descending axonal pathways closely resemble those found in frogs, reptiles, and birds. Their role in visual and visuomotor functions is discussed.     

The retinohypothalamic tract originates from a distinct subset of retinal ganglion cells

The retinal ganglion cells giving rise to retinohypothalamic projections in the rat were identified using retrograde transport of horseradish peroxidase (HRP) or FluoroGold injected into the suprachiasmatic nucleus (SCN), and using transneuronal transport of the Bartha strain of the swine herpesvirus (PRV-Bartha). When PRV-Bartha is injected into one eye, it is taken up by retinal ganglion cells, replicated, transported to axon terminals in the SCN, and released. There the virus may take one, or both, of two paths to retinal ganglion cells in the contralateral eye: 1) uptake by SCN neurons, replication, and release from the neurons with uptake and retrograde transport in retinal afferents originating in the contralateral retina; 2) transneuronal passage through axo-axonic appositions between retinal afferents in the SCN with subsequent retrograde transport of virus to the contralateral retina. The ganglion cells thus labeled are a homogeneous population of small neurons (mean diameter, 12.8 +/- 2.2 microns and mean area, 81.8 +/- 21.8 microns 2) with sparsely branching dendrites that are widely distributed over the retina. This population is best identified when virus labeling of retinal projections in areas beyond the hypothalamus is eliminated by lateral geniculate lesions that transect the optic tract at its entry into the geniculate complex. The same population is labeled with retrograde tracers but, with both HRP and FluoroGold, other ganglion cells are labeled, presumably from uptake by fibers of passage, indicating that the virus is a more reliable marker for ganglion cells giving rise to retinohypothalamic projections. The ganglion cells identified correspond to a subset of type III, or W, cells.     

Flank-marking behavior and the neural distribution of vasopressin innervation in golden hamsters with suprachiasmatic lesions

In golden hamsters, microinjections of arginine-vasopressin (AVP) within the anterior hypothalamus trigger a stereotyped scent-marking behavior, flank marking. Our experiment was carried out to test the contribution of AVP neurons within the suprachiasmatic nucleus (SCN) in the control of this behavior. Our results suggest that the SCN does not contribute to flank-marking behavior. Whereas SCN lesions disrupted circadian rhythms of wheel running, the same lesions did not disrupt flank-marking. The results also suggest that neurons located outside the SCN contribute significantly to the vasopressinergic innervation of the brain and the expression of AVP-dependent behaviors, such as flank-marking behavior. Although AVP-immunoreactive fibers were severely (ca. 95%) depleted from several forebrain areas in SCN-lesioned hamsters, the effect of the lesions was much more limited within the forebrain areas involved in flank-marking behavior as well as within the midbrain and hindbrain.     

Interconnections among nuclei of the subcortical visual shell: the intergeniculate leaflet is a major constituent of the hamster subcortical visual system

The intergeniculate leaflet (IGL), a major constituent of the circadian visual system, is one of 12 retinorecipient nuclei forming a "subcortical visual shell" overlying the diencephalic-mesencephalic border. The present investigation evaluated IGL connections with nuclei of the subcortical visual shell and determined the extent of interconnectivity between these nuclei. Male hamsters received stereotaxic, iontophoretic injections of the retrograde tracer, cholera toxin beta fragment, or the anterograde tracer, Phaseolus vulgaris-leucoagglutin, into nuclei of the pretectum (medial, commissural, posterior, olivary, anterior, nucleus of the optic tract, posterior limitans), into the superior colliculus, or into the visual thalamic nuclei (lateral posterior, dorsal lateral geniculate, intergeniculate leaflet, ventral lateral geniculate). Retrogradely labeled cell bodies identified nuclei with afferents projecting to the site of injection, whereas the presence of anterogradely labeled fibers with terminals revealed brain nuclei targeted by neurons at the site of injection. The IGL projects bilaterally to all nuclei of the visual shell except the lateral posterior and dorsal lateral geniculate nuclei. The IGL also has afferents from the same set of nuclei, except the nucleus of the optic tract. The extensive bilateral efferent projections distinguish IGL from the ventral lateral geniculate nucleus. The superior colliculus, commissural pretectal, olivary pretectal, and posterior pretectal nuclei also project bilaterally to the majority of subcortical visual nuclei. The IGL has a well-established role in circadian rhythm regulation, but there is as yet no known function for it in the larger context of the subcortical visual system, much of which is involved in oculomotor control.     

An alternate pathway for visual signal integration into the hypothalamo-pituitary axis: retinorecipient intergeniculate neurons project to various regions of the hypothalamus and innervate neuroendocrine cells including those producing dopamine

Using tract tracing and immunocytochemistry, this study explored the connectivity between lateral geniculate efferents and neurons of the hypothalamus, including those producing dopamine, that have direct access to fenestrated capillaries. It was also determined whether the intergeniculate neurons that give rise to hypothalamic projections are targeted by retinal axons. Within the hypothalamus, Phaseolus vulgaris leucoagglutinin-labeled, lateral geniculate efferents were observed in the suprachiasmatic nucleus, subparaventricular area, periventricular nuclei, medial preoptic areas, and between the arcuate and ventromedial nuclei. In these sites, intergeniculate efferents contacted populations of neurons that were retrogradely labeled from fenestrated capillaries by the intraperitoneal injection of fluorogold. Hypothalamic dopamine neurons, a population of which was neuroendocrine, were also synaptic targets of lateral geniculate efferents. After injection of the retrograde tracer fluorogold into these hypothalamic projection sites in parallel with bilateral enucleation, retrogradely labeled perikarya were restricted to the intergeniculate leaflet. All of the labeled perikarya contained infolded nuclei, and their distal dendrites were frequently found to be contacted by degenerated, retinal fibers. This study provides morphological evidence for a signaling pathway from the retina through the intergeniculate leaflet to hypothalamic cells that participate in neuroendocrine regulations. These observations raise the possibility that visual signals independent of the circadian clock may also influence the hypothalamo-pituitary axis. In light of the overlapping distribution of intergeniculate and suprachiasmatic efferents in the hypothalamus and their similar relationship with neuroendocrine cells, it is suggested that integration of circadian and visual signals can occur outside of the suprachiasmatic nucleus to regulate endocrine rhythms.