Jeferson S. Cavalcante

Retinohypothalamic projections in the common marmoset (Callithrix jacchus): A study using cholera toxin subunit B.

Retinal projections in vertebrates reach the primary visual, accessory optic, and circadian timing structures. The central feature of the circadian timing system is the principal circadian pacemaker, the suprachiasmatic nucleus (SCN) of the hypothalamus. The direct projections from the retina to the SCN are considered the entrainment pathway of the circadian timing system. In this study, unilateral intravitreal injections of cholera toxin subunit B were used to trace the retinal projections to the marmoset hypothalamus. The retinohypothalamic tract reaches the ventral suprachiasmatic nucleus bilaterally, as anticipated from previous studies. However, labeled fibers were found in several other hypothalamic regions, such as the medial and lateral preoptic areas, supraoptic nucleus, anterior and lateral hypothalamic areas, retrochiasmatic area, and subparaventricular zone. These results reveal new aspects of retinohypothalamic projection in primates and are discussed in terms of their implications for circadian as well as noncircadian control systems. J. Comp. Neurol. 415:393–403, 1999.

Cognitive, motor and tyrosine hydroxylase temporal impairment in a model of parkinsonism induced by reserpine

Studies have suggested that cognitive deficits can precede motor alterations in Parkinsons disease (PD). However, in general, classic animal models are based on severe motor impairment after one single administration of neurotoxins, and thereby do not express the progressive nature of the pathology. A previous study showed that the repeated administration with a low dose (0.1mg/kg) of the monoamine depleting agent reserpine induces a gradual appearance of motor signs of pharmacological parkinsonism in rats. Here, we showed this repeated treatment with reserpine induced a memory impairment (evaluated by the novel object recognition task) before the gradual appearance of the motor signs. Additionally, these alterations were accompanied by decreased tyrosine hydroxylase (TH) striatal levels and reduced number of TH+ cells in substantia nigra pars compacta (SNpc). After 30 days without treatment, reserpine-treated animals showed normal levels of striatal TH, partial recovery of TH+ cells in SNpc, recovery of motor function, but not reversal of the memory impairment. Furthermore, the motor alterations were statistically correlated with decreased TH levels (GD, CA1, PFC and DS) and number of TH+ cells (SNpc and VTA) in the brain. Thus, we extended previous results showing that the gradual appearance of motor impairment induced by repeated treatment with a low dose of reserpine is preceded by short-term memory impairment, as well as accompanied by neurochemical alterations compatible with the pathology of PD.

Basolateral amygdala inactivation impairs learned (but not innate) fear response in rats

Numerous studies have suggested that the amygdala is involved in the formation of aversive memories, but the possibility that this structure is merely related to any kind of fear sensation or response could not be ruled out in previous studies. The present study investigated the effects of bilateral inactivation of the amygdaloid complex in rats tested in the plus-maze discriminative avoidance task. This task concomitantly evaluates aversive memory (by discrimination of the two enclosed arms) and innate fear (by open-arm exploration). Wistar rats (3-5 months-old) were implanted with bilateral guide cannulae into basolateral amygdala. After surgery, all subjects were given 1 week to recover before behavioral experiments. Afterwards, in experiment 1, 15 min prior to training, 0.5 μl of saline or muscimol (1 mg/ml) was infused in each side via microinjection needles. In experiment 2 the animals received injections immediately after the training session and in experiment 3 rats were injected prior to testing session (24 h after training). The main results showed that (1) pre-training muscimol prevented memory retention (evaluated by aversive arm exploration in the test session), but did not alter innate fear (evaluated by percent time in open arms); (2) post-training muscimol impaired consolidation, inducing increased percent in aversive arm exploration in the test session and (3) pre-testing muscimol did not affect retrieval (evaluated by aversive enclosed arm exploration in the test session). The results suggest that amygdala inactivation specifically modulated the learning of the aversive task, excluding a possible secondary effect of amygdala inactivation on general fear responses. Additionally, our data corroborate the hypothesis that basolateral amygdala is not the specific site of storage of aversive memories, since retention of the previously learned task was not affected by pre-testing inactivation.

Evolutionary History of the PER3 Variable Number of Tandem Repeats (VNTR): Idiosyncratic Aspect of Primate Molecular Circadian Clock

The PER3 gene is one of the clock genes, which function in the core mammalian molecular circadian system. A variable number of tandem repeats (VNTR) locus in the 18th exon of this gene has been strongly associated to circadian rhythm phenotypes and sleep organization in humans, but it has not been identified in other mammals except primates. To better understand the evolution and the placement of the PER3 VNTR in a phylogenetical context, the present study enlarges the investigation about the presence and the structure of this variable region in a large sample of primate species and other mammals. The analysis of the results has revealed that the PER3 VNTR occurs exclusively in simiiforme primates and that the number of copies of the primitive unit ranges from 2 to 11 across different primate species. Two transposable elements surrounding the 18th exon of PER3 were found in primates with published genome sequences, including the tarsiiforme Tarsius syrichta, which lacks the VNTR. These results suggest that this VNTR may have evolved in a common ancestor of the simiiforme branch and that the evolutionary copy number differentiation of this VNTR may be associated with primate simiiformes sleep and circadian phenotype patterns.

Retinal projections to the midline and intralaminar thalamic nuclei in the common marmoset (Callithrix jacchus)

In this study, we report the identification of a hitherto not reported direct retinal projection to midline and intralaminar thalamic nuclei in the marmoset brain. After unilateral intravitreal injections of cholera toxin subunit B (CTb), anterogradely transported CTb-immunoreactive fibers and presumptive terminals were seen in the following thalamic midline nuclei: paraventricular, rhomboid, interanteromedial, and reuniens, and thalamic intralaminar nuclei: central medial, central lateral, central dorsal, and parafascicular. Studies employing sensitive tracers in other primate species are needed in order to verify the possible universality of these projections. Some of the possible functional correlates of the present data are briefly discussed. The present results may contribute to the elucidation of the anatomical substrate of the functionally demonstrated involvement of this midline/intralaminar thalamic nuclear complex in several domains that include arousal and awareness, besides specific cognitive, sensory, and motor functions.

Immunocytochemical characterization of the pregeniculate nucleus and distribution of retinal and neuropeptide Y terminals in the suprachiasmatic nucleus of the Cebus monkey

Circadian rhythms generated by the suprachiasmatic nucleus (SCN) are modulated by photic and non-photic stimuli. In rodents, direct photic stimuli reach the SCN mainly through the retinohypothalamic tract (RHT), whereas indirect photic stimuli are mainly conveyed by the geniculohypothalamic tract (GHT). In rodents, retinal cells form a pathway that reaches the intergeniculate leaflet (IGL) where they establish synapses with neurons that express neuropeptide Y (NPY), hence forming the GHT projecting to the SCN. In contrast to the RHT, which has been well described in primates, data regarding the presence or absence of the IGL and GHT in primates are contradictory. Some studies have suggested that an area of the pregeniculate nucleus (PGN) of primates might be homologous to the IGL of rodents, but additional anatomical and functional studies on primate species are necessary to confirm this hypothesis. Therefore, this study investigated the main histochemical characteristics of the PGN and the possible existence of the GHT in the SCN of the primate Cebus, comparing the distribution of NPY immunoreactivity, serotonin (5-HT) immunoreactivity and retinal terminal fibers in these two structures. The results show that a collection of cell bodies containing NPY and serotonergic immunoreactivity and retinal innervations are present within a zone that might be homologous to the IGL of rodents. The SCN also receives dense retinal innervations and we observed an atypical distribution of NPY- and 5-HT-immunoreactive fibers without regionalization in the ventral part of the nucleus as described for other species. These data may reflect morphological differences in the structures involved in the regulation of circadian rhythms among species and support the hypothesis that the GHT is present in some higher primates (diurnal animals).

Sciatic nerve grafting and inoculation of FGF-2 promotes improvement of motor behavior and fiber regrowth in rats with spinal cord transection

PURPOSE Failure of severed adult central nervous system (CNS) axons to regenerate could be attributed with a reduced intrinsic growing capacity. Severe spinal cord injury is frequently associated with a permanent loss of function because the surviving neurons are impaired to regrow their fibers and to reestablish functional contacts. Peripheral nerves are known as good substrate for bridging CNS trauma with neurotrophic factor addition. We evaluated whether fibroblastic growth factor 2 (FGF-2) placed in a gap promoted by complete transection of the spinal cord may increase the ability of sciatic nerve graft to enhance motor recovery and fibers regrow. METHODS We used a complete spinal cord transection model. Rats received a 4 mm-long gap at low thoracic level and were repaired with saline (control) or fragment of the sciatic nerve (Nerve) or FGF-2 was added to nerve fragment (Nerve+FGF-2) to the grafts immediately after complete transection. The hind limbs performance was evaluated weekly for 8 weeks by using motor behavior score (BBB) and sensorimotor tests-linked to the combined behavior score (CBS), which indicate the degree of the motor improvement and the percentage of functional deficit, respectively. Neuronal plasticity were evaluated at the epicenter of the injury using MAP-2 and GAP-43 expression. RESULTS Spinal cord treatment with sciatic nerve and sciatic nerve plus FGF-2 allowed recovery of hind limb movements compared to control, manifested by significantly higher behavioral scores. Higher amounts of MAP-2 and GAP-43 immunoreactive fibers were found in the epicenter of the graft when FGF-2 was added. CONCLUSIONS FGF-2 added to the nerve graft favored the motor recovery and fiber regrowth. Thus, these results encourage us to explore autologous transplantation as a novel and promising cell therapy for treatment of spinal cord lesion.

The suprachiasmatic nucleus and the intergeniculate leaflet in the rock cavy (Kerodon rupestris): Retinal projections and immunohistochemical characterization

In this study, two circadian related centers, the suprachiasmatic nucleus (SCN) and the intergeniculate leaflet (IGL) were evaluated in respect to their cytoarchitecture, retinal afferents and chemical content of major cells and axon terminals in the rock cavy (Kerodon rupestris), a Brazilian rodent species. The rock cavy SCN is innervated in its ventral portion by terminals from the predominantly contralateral retina. It also contains vasopressin, vasoactive intestinal polypeptide and glutamic acid decarboxilase immunoreactive cell bodies and neuropeptide Y, serotonin and enkephalin immunopositive fibers and terminals and is marked by intense glial fibrillary acidic protein immunoreactivity. The IGL receives a predominantly contralateral retinal projection, contains neuropeptide Y and nitric oxide synthase-producing neurons and enkephalin immunopositive terminals and is characterized by dense GFAP immunoreactivity. This is the first report examining the neural circadian system in a crepuscular rodent species for which circadian properties have been described. The results are discussed comparing with what has been described for other species and in the context of the functional significance of these centers.

Retinal projections to the thalamic paraventricular nucleus in the rock cavy (Kerodon rupestris)

The thalamic paraventricular nucleus (PVT) receives afferents from numerous brain areas, including the hypothalamic suprachiasmatic nucleus (SCN), considered to be the major circadian pacemaker. The PVT also sends projections to the SCN, limbic system centers and some nuclei involved in the control of the Sleep-Wake cycle. In this study, we report the identification of a hitherto not reported direct retinal projection to the PVT of the rock cavy, a typical rodent species of the northeast region of Brazil. After unilateral intravitreal injections of cholera toxin subunit B (CTb), anterogradely transported CTb-immunoreactive fibers and presumptive terminals were seen in the PVT. Some possible functional correlates of the present data are briefly discussed, including the role of the PVT in the modulation of the circadian rhythms by considering the reciprocal connections between the PVT and the SCN. The present work is the first to show a direct retinal projection to the PVT of a rodent and may contribute to elucidate the anatomical substrate of the functionally demonstrated involvement of this midline thalamic nucleus in the modulation of the circadian timing system.

Mediodorsal thalamic nucleus receives a direct retinal input in marmoset monkey (Callithrix jacchus): a subunit B cholera toxin study.

The mediodorsal thalamic nucleus is a prominent nucleus in the thalamus, positioned lateral to the midline nuclei and medial to the intralaminar thalamic complex in the dorsal thalamus. Several studies identify the mediodorsal thalamic nucleus as a key structure in learning and memory, as well as in emotional mechanisms and alertness due to reciprocal connections with the limbic system and prefrontal cortex. Fibers from the retina to the mediodorsal thalamic nucleus have recently been described for the first time in a crepuscular rodent, suggesting a possible regulation of the mediodorsal thalamic nucleus by visual activity. The present study shows retinal afferents in the mediodorsal thalamic nucleus of a new world primate, the marmoset (Callithrix jacchus), using B subunit of cholera toxin (CTb) as an anterograde tracer. A small population of labeled retinofugal axonal arborizations is consistently labeled in small domains of the medial and lateral periphery of the caudal half of the mediodorsal nucleus. Retinal projections in the mediodorsal thalamic nucleus are exclusively contralateral and the morphology of the afferent endings was examined. Although the functional significance of this projection remains unknown, this retina-mediodorsal thalamic nucleus pathway may be involved in a wide possibility of functional implications.