Preoptic Nucleus Research Articles

β-Lapachone Exerts Hypnotic Effects via Adenosine A1 Receptor in Mice.

Sleep is one of the most essential physiological phenomena for maintaining health. Sleep disturbances, such as insomnia, are often accompanied by psychiatric or physical conditions such as impaired attention, anxiety, and stress. Medication used to treat insomnia have concerns about potential side effects with long-term use, so interest in the use of alternative medicine is increasing. In this study, we investigated the hypnotic effects of β-lapachone (β-Lap), a natural naphthoquinone compound, using pentobarbital-induced sleep test, immunohistochemistry, real-time PCR, and western blot in mice. Our results indicated that β-Lap exerts a significant hypnotic effect by showing a decrease in sleep onset latency and an increase in total sleep time in pentobarbital-induced sleep model. The results of c-Fos immunostaining showed that β-Lap decreased neuronal activity in the basal forebrain and lateral hypothalamus, which are wakefulness-promoting brain regions, while increasing in the ventrolateral preoptic nucleus, a sleep-promoting region; all these effects were significantly abolished by 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), an adenosine A1 receptor (A1R) antagonist. Western blot analysis showed that β-Lap increased extracellular signalregulated kinase phosphorylation and nuclear factor-kappa B translocation from the cytoplasm to the nucleus; these effects were inhibited by DPCPX. Additionally, β-Lap increased the mRNA levels of A1R. Taken together, these results suggest that β-Lap exerts hypnotic effects, potentially through A1R.

Whole-brain mapping in adult zebrafish and identification of a novel tank test functional connectome.

Identifying general principles of brain function requires the study of structure-function relationships in a variety of species. Zebrafish have recently gained prominence as a model organism in neuroscience, yielding important insights into vertebrate brain function. Although methods have been developed for mapping neural activity in larval animals, we lack similar techniques for adult zebrafish that have the advantage of a fully developed neuroanatomy and larger behavioral repertoire. Here, we describe a pipeline built around open-source tools for whole-brain activity mapping in freely swimming adult zebrafish. Our pipeline combines recent advances in histology, microscopy, and machine learning to capture cfos activity across the entirety of the adult brain. Images captured using light-sheet microscopy are registered to the recently created adult zebrafish brain atlas (AZBA) for automated segmentation using advanced normalization tools (ANTs). We used our pipeline to measure brain activity after zebrafish were subject to the novel tank test. We found that cfos levels peaked 15 minutes following behavior and that several regions containing serotoninergic, dopaminergic, noradrenergic, and cholinergic neurons were active during exploration. Finally, we generated a novel tank test functional connectome. Functional network analysis revealed that several regions of the medial ventral telencephalon form a cohesive sub-network during exploration. We also found that the anterior portion of the parvocellular preoptic nucleus (PPa) serves as a key connection between the ventral telencephalon and many other parts of the brain. Taken together, our work enables whole-brain activity mapping in adult zebrafish for the first time while providing insight into neural basis for the novel tank test.

Distinct populations of lateral preoptic nucleus neurons jointly contribute to depressive-like behaviors through divergent projections in male mice

The lateral preoptic area (LPO) is a component of the hypothalamus involved in various physiological functions including sleep-wakefulness transition, thermoregulation, and water-salt balance. In this study, we discovered that distinct LPO excitatory neurons project separately to the aversive processing center lateral habenula (LHb) and the reward processing hub ventral tegmental area (VTA). Following chronic restraint stress (CRS), the LHb-projecting and VTA-projecting LPO neurons exhibited increased and decreased neuronal activities, respectively. Optogenetic activation of LHb-projecting LPO excitatory neurons and LPO excitatory neuronal terminals within LHb evoked aversion and avoidance behaviors, while activation of VTA-projecting LPO excitatory neurons and LPO excitatory neuronal terminals within VTA produced preference and exploratory behaviors in mice. Furthermore, either optogenetic inhibition of LHb-projecting LPO excitatory neurons or activation of VTA-projecting LPO excitatory neurons during CRS effectively prevented the development of depressive-like behaviors. Our study unveils, for the first-time, divergent pathways originating from LPO that regulate opposite affective states in mice and implicates that an imbalance of their activities could lead to depressive-like behaviors. These circuitries represent promising therapeutic targets to relieve emotional dysfunctions in neuropsychiatric disorders.

Spontaneous and evoked angiotensin II sniffer cell activity in the lamina terminalis in vitro.

Angiotensin II (ANG II) has been shown to have central nervous system effects. Although tissue renin-angiotensin systems (RAS) have been demonstrated in multiple tissues, the existence of a brain RAS is still a matter of debate. These studies test for angiotensin release from brain slices prepared from adult male Sprague-Dawley rats and male and female renin knock-out rats using Chinese hamster ovary cells modified to express both the angiotensin II type 1 receptor and a fluorescent calcium indicator. Sniffer cells were placed on the slices and calcium transients were measured from those located on or adjacent to the median preoptic nucleus with and without stimulation of the subfornical organ. Bath application of tetrodotoxin (1 µM) significantly attenuated spontaneous events while abolishing evoked sniffer cell activity. Bath application of dl-AP4 (10 µM, glutamatergic antagonist) did not affect either spontaneous or evoked release. Incubating the slices with fluorocitrate to inactive astrocytes did not influence sniffer cell activity in the MnPO. Pharmacological experiments indicate that ANG II release is largely both renin (aliskiren 10 µM) and ACE-1 (captopril 100 µM) dependent. However, experiments with brain slices prepared from male and female Renin knock-out rats suggest that alternative synthetic pathways may exist. Finally, these studies demonstrate that increases in ANG II release are observed following 7 days of chronic intermittent hypoxia. These studies suggest the existence of a tissue-specific RAS in the brain that involves canonical and alternative ANG II synthetic pathways and is upregulated in an animal model of sleep apnea.NEW & NOTEWORTHY These studies used Chinese hamster ovary cells that were cloned to express an angiotensin receptor (At1ra) and a calcium indicator (R-GECO) to detect the release of angiotensin from brain slices containing the lamina terminalis of rats. Some of the experiments use tissue from renin knockout rats. The results support the existence of an angiotensin system in the brain that may involve alternative synthetic pathways and is upregulated by intermittent hypoxia.

Sedum kamtschaticum Exerts Hypnotic Effects via the Adenosine A2A Receptor in Mice.

Insomnia is a common sleep disorder with significant societal and economic impacts. Current pharmacotherapies for insomnia are often accompanied by side effects, necessitating the development of new therapeutic drugs. In this study, the hypnotic effects and mechanisms of Sedum kamtschaticum 30% ethanol extract (ESK) and one of its active compounds, myricitrin, were investigated using pentobarbital-induced sleep experiments, immunohistochemistry (IHC), receptor binding assays, and enzyme-linked immunosorbent assay (ELISA). The pentobarbital-induced sleep experiments revealed that ESK and myricitrin reduced sleep latency and prolonged total sleep time in a dose-dependent manner. Based on c-Fos immunostaining, ESK, and myricitrin enhanced the GABAergic neural activity in sleep-promoting ventrolateral preoptic nucleus (VLPO) GABAergic. By measuring the level of GABA released from VLPO GABAergic neurons, ESK and myricitrin were found to increase GABA release in the hypothalamus. These effects were significantly inhibited by SCH. Moreover, ESK exhibited a concentration-dependent binding affinity for the adenosine A2A receptors (A2AR). In conclusion, ESK and myricitrin have hypnotic effects, and their underlying mechanisms may be related to the activation of A2AR.

Hypnotic effect of AR-001 through adenosine A1 receptor

Insomnia is one of the most common sleep disorders, affecting 10–15% of the global population. Because classical remedies used to treat insomnia have various side effects, new therapeutics for insomnia are attracting attention. In the present study, we found that N2-Ethyl-N4-(furan-2-ylmethyl) quinazoline-2,4-diamine (AR-001) has adenosine A1 receptor agonistic activity and exhibits hypnotic efficacy by decreasing sleep onset latency and increasing total sleep time in a pentobarbital-induced sleep model. This hypnotic effect of AR-001 was significantly inhibited by the adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX). As a result of immunohistochemistry, AR-001 was shown to increase neural activity in the sleep-promoting region, ventrolateral preoptic nucleus (VLPO), and decrease neural activity in the wake-promoting region, basal forebrain (BF), and lateral hypothalamus (LH), and that these effects of AR-001 were significantly inhibited by DPCPX treatment. In addition, AR-001 increased adenosine A1 receptor mRNA levels in the hypothalamus. In conclusion, this study suggests that AR-001 has a hypnotic effect, at least partially, through adenosine A1 receptor and may have therapeutic potential for insomnia.

Role of the Locus Coeruleus Arousal Promoting Neurons in Maintaining Brain Criticality across the Sleep-Wake Cycle.

Sleep control depends on a delicate interplay among brain regions. This generates a complex temporal architecture with numerous sleep-stage transitions and intermittent fluctuations to micro-states and brief arousals. These temporal dynamics exhibit hallmarks of criticality, suggesting that tuning to criticality is essential for spontaneous sleep-stage and arousal transitions. However, how the brain maintains criticality remains not understood. Here, we investigate θ- and δ-burst dynamics during the sleep-wake cycle of rats (Sprague-Dawley, adult male) with lesion in the wake-promoting locus coeruleus (LC). We show that, in control rats, θ- and δ-bursts exhibit power-law (θ-bursts, active phase) and exponential-like (δ-bursts, quiescent phase) duration distributions, as well as power-law long-range temporal correlations (LRTCs)-typical of non-equilibrium systems self-organizing at criticality. Furthermore, consecutive θ- and δ-bursts durations are characterized by anti-correlated coupling, indicating a new class of self-organized criticality that emerges from underlying feedback between neuronal populations and brain areas involved in generating arousals and sleep states. In contrast, we uncover that LC lesion leads to alteration of θ- and δ-burst critical features, with change in duration distributions and correlation properties, and increase in θ-δ coupling. Notably, these LC-lesion effects are opposite to those observed for lesions in the sleep-promoting ventrolateral preoptic (VLPO) nucleus. Our findings indicate that critical dynamics of θ- and δ-bursts arise from a balanced interplay of LC and VLPO, which maintains brain tuning to criticality across the sleep-wake cycle-a non-equilibrium behavior in sleep micro-architecture at short timescales that coexists with large-scale sleep-wake homeostasis.

Regulation of sleep by astrocytes in the hypothalamic ventrolateral preoptic nucleus.

Regulation of sleep by astrocytes in the hypothalamic ventrolateral preoptic nucleus.

Ovariectomy Increases Blood Pressure and FosB Staining in the Central Autonomic Control Regions Independent of Chronic Intermittent Hypoxia

Chronic intermittent hypoxia (CIH) experimentally mimics the recurrent hypoxia characteristic of sleep apnea, and it induces sustained hypertension in male and ovariectomized (OVX) female rats but not intact females. While central autonomic control regions are known to mediate CIH-induced hypertension in males, their contribution to CIH hypertension in OVX females remains unclear. We tested the hypothesis that 7 days of CIH increases mean arterial pressure (MAP) and FosB staining in central autonomic regions in OVX but not intact female rats. Adult female Sprague Dawley gonadally intact (INT) or OVX rats were exposed to 7 days of CIH (10% O2 and 21% O2 cycle, every 6 mins, 8h/day) or continuous normoxia (CON). MAP was monitored by radiotelemetry. On day 8, the rats were sacrificed, and their brains were collected and processed for FosB immunohistochemistry. During the light/sleep phase, CIH increased MAP in INT females (n=7; 3.4 ± 0.4 mmHg) as compared to CON (n=6, ΔMAP 1.4 ± 0.3 mmHg; P=0.01 CON vs CIH). This difference was not observed during the dark/active phase (CON ΔMAP -1.7 ± 0.4 mmHg, CIH 0.5 ± 0.3 mmHg, P=0.07). In OVX females, CIH was not associated with significant increases in MAP during the light/sleep phase (CON (n=6) ΔMAP 0.3 ± 0.4 mmHg, CIH (n = 6) 2.1 ± 0.3 mmHg, P=0.10). During the dark phase, MAP was significantly increased in CON OVX and CIH OVX compared to their respective baselines but there were no differences between groups (CON ΔMAP 2.4 ± 0.3 mmHg, CIH 2.2 ± 0.4 mmHg, P=0.0002 vs baseline, P=0.8 CON vs CIH). OVX rats had higher baseline FosB staining in the median preoptic nucleus compared to intact female rats independent of CIH exposure (INT: CON (n = 6) 13 ± 3 cells/section, CIH (n = 6) 14 ± 2; OVX: CON (n = 6) 21 ± 3, CIH (n = 6) 23 ± 2, INT CON vs OVX CON, P < 0.05, INT CIH vs OVX CIH P < 0.05). Compared to CON, CIH significantly increased the number of FosB immunoreactive cells in the caudal nucleus of the solitary tract in INT but not OVX females (INT: CON, 7 ± 1 cells/section, CIH 11 ± 1; OVX CON 6 ± 1, CIH n=6, 9 ± 1; INT CON vs CIH, P=0.02; OVX CON vs CIH, P = 0.09). In the rostral ventrolateral medulla, there were no significant differences among any of the groups (INT: CON n=4, 19 ± 2 cells/section, CIH n=4, 20 ± 3; OVX: CON n=6, 18 ± 2, CIH n=6, 21 ± 2, all P>0.05). In OVX females, CIH tended to increase MAP during both the light and dark phases. CON OVX females showed a trend for increased MAP during the dark phase, independent of CIH. CIH was not associated with increased FosB staining in either the median preoptic nucleus or the rostral ventrolateral medulla of OVX females This suggests that changes in MAP observed in CIH OVX females may not involve the same autonomic control regions as male rats exposed to CIH. Supported by RO1 HL155977. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Role of the median preoptic nucleus in the development of chronic deoxycorticosterone acetate (DOCA)-salt hypertension.

We have previously reported that the subfornical organ (SFO) does not contribute to the chronic hypertensive response to DOCA-salt in rats, and yet the organum vasculosum of the lamina terminalis (OVLT) plays a significant role in the development of deoxycorticosterone acetate (DOCA)-salt hypertension. Since efferent fibers of the OVLT project to and through the median preoptic nucleus (MnPO), the present study was designed to test the hypothesis that the MnPO is necessary for DOCA-salt hypertension in the rat. Male Sprague-Dawley rats underwent SHAM (MnPOsham; n = 5) or electrolytic lesion of the MnPO (MnPOx; n = 7) followed by subsequent unilateral nephrectomy and telemetry instrumentation. After recovery and during the experimental protocol, rats consumed a 0.1% NaCl diet and 0.9% NaCl drinking solution. Mean arterial pressure (MAP) was recorded telemetrically 5 days before and 21 days after DOCA implantation (100 mg/rat; SQ). The chronic pressor response to DOCA was attenuated in MnPOx rats by Day 11 of treatment and continued such that MAP increased 25 ± 3 mmHg in MnPOsham rats by Day 21 of DOCA compared to 14 ± 3 mmHg in MnPOx rats. These results support the hypothesis that the MnPO is an important brain site of action and necessary for the full development of DOCA-salt hypertension in the rat.

Effects of OVLT or MnPO lesion on the gut microbiome

Hypertension is a multifaceted condition affecting 700 million people worldwide, and despite decades of study, several aspects of its complex pathophysiology remain unclear. Changes in the gut microbiome have been associated with hypertension, though the cause and effect are not well defined. Historically, circumventricular organs in the brain, such as the organum vasculosum of the lamina terminalis (OVLT), and its central downstream nucleus the median preoptic nucleus (MnPO) have been shown to play a role in the pathogenesis of hypertension. Studies from our lab have demonstrated roles of both the OVLT and MnPO in the pathogenesis of angiotensin II mediated hypertension. Furthermore, we have previously reported that DOCA-salt treated OVLT lesioned rats had attenuated hypertension, and a gut microbial population more closely resembling normal controls, compared to sham lesioned rats. Lastly, we have recently shown the MnPO to be necessary in mediating DOCA-salt hypertension in the rat. Because data suggests changes in the gut microbiome occurred during the attenuated hypertensive response in lesioned animals, it is still unclear whether these changes occurred due to and shortly after the lesion, or subsequently during the hypertensive stimulus in lesioned animals. Therefore, the goal of this study was to further define the relationship between an OVLT or MnPO lesion and consequent changes in the gut microbiome without any hypertensive stimulus. Male Sprague Dawley rats underwent electrolytic lesion or sham operation using stereotaxic surgery with predetermined coordinates and settings. After 7 days, brain and regional gut tissues were harvested for analysis. Gut microbial genomic data from lesioned and sham rats were analyzed, and PCoA analysis showed statistically significant differences in microbiome composition between OVLT lesioned and sham rats in most gut regions. Generally, Shannon diversity analysis revealed sham rats had a more diverse population than OVLT lesioned rats, though both had a relatively similar spread of species abundance. Additionally, there was a common trend of higher relative abundance of Ligilactobacillus in lesioned rats for all four gut regions. In conclusion, the OVLT lesion itself has effects on the diversity of the gut microbiome, which suggest an OVLT-gut microbiome link that may contribute to attenuating hypertension. This study was supported by University of Minnesota Grant-In-Aid of Research, Artistry and Scholarship #546839 and a University of Minnesota Lillehei Heart Institute Collaborative Pilot Grant. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Characterisation of a Novel Osmosensitive Pathway from the Supraoptic Nucleus to the Median Preoptic Nucleus

The Supraoptic Nucleus (SON) is well regarded for the neuroendocrine release of vasopressin and oxytocin through the posterior pituitary gland. Evidence also exists for extra-pituitary efferent projections of SON neurons, mainly collaterals to the lateral hypothalamus and central amygdala. The objective of this study was to examine the existence of a central SON projection to the median preoptic nucleus (MnPO) of the hypothalamus. Microinjections of the retrograde tracer Cholera Toxin B Subunit (CTb) into the MnPO of male Wistar rats (n = 3) retrogradely labelled SON neurons. Fluorescence immunohistochemistry revealed equal involvement of vasopressin (AVP) and oxytocin (OXT) — producing SON neurons (n = 107 AVP+/CTb+, n = 110 OXT+/ CTb+, n = 417 total CTb+ neurons). This major collateral projection was also examined in an animal model of Polycystic Kidney Disease, the Lewis Polycystic Kidney (LPK) rat (n = 3 animals; n = 104 AVP+/CTb+, n = 41 OXT+/CTb+, n = 372 total CTb+ neurons) and its healthy Lewis control (n = 3 animals; n = 115 AVP+/CTb+, n = 100 OXT+/CTb+, n = 430 total CTb+ neurons). Although the function of this projection is currently unclear, preliminary data in a subset of LPKs shows these SON neurons with MnPO collateral projections are activated by osmotic stressors (e.g., hypertonic saline). In summary, we report a major collateral projection from the SON to MnPO that is activated by osmotic stress. Given the well-known role of the MnPO in drinking behaviour, we suggest this SON-MnPO circuit may be involved in the coordination of fluid regulation and drinking behaviour. This project was funded by Macquarie University. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

0046 A Mechanism by Which Estradiol (E2) Regulates Adenosinergic Signaling in the Median Preoptic Nucleus (MnPO)

0046 A Mechanism by Which Estradiol (E2) Regulates Adenosinergic Signaling in the Median Preoptic Nucleus (MnPO)

0043 The Role of Median Preoptic Nucleus Astrocytes in Mediating E2’s Effects on Sleep-Wake Behavior

0043 The Role of Median Preoptic Nucleus Astrocytes in Mediating E2’s Effects on Sleep-Wake Behavior

Paracrine and endocrine pathways of natriuretic peptides assessed by ligand-receptor mapping in the Japanese eel brain.

The natriuretic peptide (NP) family consists of cardiac NPs (ANP, BNP, and VNP) and brain NPs (CNPs) in teleosts. In addition to CNP1-4, a paralogue of CNP4 (named CNP4b) was recently discovered in basal teleosts including Japanese eel. Mammals have lost most Cnps during the evolution, but teleost cnps were conserved and diversified, suggesting that CNPs are important hormones for maintaining brain functions in teleost. The present study evaluated the potency of each Japanese eel CNP to their NP receptors (NPR-A, NPR-B, NPR-C, and NPR-D) overexpressed in CHO cells. A comprehensive brain map of cnps- and nprs-expressing neurons in Japanese eel was constructed by integrating the localization results obtained by in situ hybridization. The result showed that CHO cells expressing NPR-A and NPR-B induced strong cGMP productions after stimulation by cardiac and brain NPs, respectively. Regarding brain distribution of cnps, cnp1 is engaged in the ventral telencephalic area and periventricular area including the parvocellular preoptic nucleus (Pp), anterior/posterior tuberal nuclei, and periventricular gray zone of the optic tectum. cnp3 is found in the habenular nucleus and prolactin cells in the pituitary. cnp4 is expressed in the ventral telencephalic area, while cnp4b is expressed in the motoneurons in the medullary area. Such CNP isoform-specific localizations suggest that function of each CNP has diverged in the eel brain. Furthermore, the Pp lacking the blood-brain barrier expressed both npra and nprb, suggesting that endocrine and paracrine NPs interplay for regulating the Pp functions in Japanese eels.

Effects of physical training on hypothalamic neuronal activation and expressions of vasopressin and oxytocin in SHR after running until fatigue.

To assess the influence of physical training on neuronal activation and hypothalamic expression of vasopressin and oxytocin in spontaneously hypertensive rats (SHR), untrained and trained normotensive rats and SHR were submitted to running until fatigue while internal body and tail temperatures were recorded. Hypothalamic c-Fos expression was evaluated in thermoregulatory centers such as the median preoptic nucleus (MnPO), medial preoptic nucleus (mPOA), paraventricular nucleus of the hypothalamus (PVN), and supraoptic nucleus (SON). The PVN and the SON were also investigated for vasopressin and oxytocin expressions. Although exercise training improved the workload performed by the animals, it was reduced in SHR and followed by increased internal body temperature due to tail vasodilation deficit. Physical training enhanced c-Fos expression in the MnPO, mPOA, and PVN of both strains, and these responses were attenuated in SHR. Vasopressin immunoreactivity in the PVN was also increased by physical training to a lesser extent in SHR. The already-reduced oxytocin expression in the PVN of SHR was increased in response to physical training. Within the SON, neuronal activation and the expressions of vasopressin and oxytocin were reduced by hypertension and unaffected by physical training. The data indicate that physical training counterbalances in part the negative effect of hypertension on hypothalamic neuronal activation elicited by exercise, as well as on the expression of vasopressin and oxytocin. These hypertension features seem to negatively influence the workload performed by SHR due to the hyperthermia derived from the inability of physical training to improve heat dissipation through skin vasodilation.

Crosstalk between the subiculum and sleep-wake regulation: A review.

The circuitry underlying the initiation, maintenance, and coordination of wakefulness, rapid eye movement sleep, and non-rapid eye movement sleep is not thoroughly understood. Sleep is thought to arise due to decreased activity in the ascending reticular arousal system, which originates in the brainstem and awakens the thalamus and cortex during wakefulness. Despite the conventional association of sleep-wake states with hippocampal rhythms, the mutual influence of the hippocampal formation in regulating vigilance states has been largely neglected. Here, we focus on the subiculum, the main output region of the hippocampal formation. The subiculum, particulary the ventral part, sends extensive monosynaptic projections to crucial regions implicated in sleep-wake regulation, including the thalamus, lateral hypothalamus, tuberomammillary nucleus, basal forebrain, ventrolateral preoptic nucleus, ventrolateral tegmental area, and suprachiasmatic nucleus. Additionally, second-order projections from the subiculum are received by the laterodorsal tegmental nucleus, locus coeruleus, and median raphe nucleus, suggesting the potential involvement of the subiculum in the regulation of the sleep-wake cycle. We also discuss alterations in the subiculum observed in individuals with sleep disorders and in sleep-deprived mice, underscoring the significance of investigating neuronal communication between the subiculum and pathways promoting both sleep and wakefulness.

Ontogenetic changes in the tyrosine hydroxylase immunoreactive preoptic area in the small-spotted catshark Scyliorhinus canicula (L., 1758) females: catecholaminergic involvement in sexual maturation.

The catecholaminergic component of the brain-pituitary-gonadal axis, which mediates the influence of external and internal stimuli on the central nervous system and gonad development in vertebrates, is largely unexplored in Chondrichthyes. We considered Scyliorhinus canicula (L., 1758) females as a model for this vertebrate's class, to assess the involvement of the catecholaminergic system of the brain in its reproduction. Along the S. canicula reproductive cycle, we characterized and evaluated differences in somata morphometry and the number of putative catecholaminergic neurons in two brain nuclei: the periventricular preoptic nucleus, hypothesized to be a positive control for ovarian development, and the suprachiasmatic nucleus, examined as a negative control. 16 S. canicula wild females were sampled and grouped in maturity stages (immature, maturing, mature, and mature egg-laying). The ovary was histologically processed for the qualitative description of maturity stages. Anti-tyrosine hydroxylase immunofluorescence was performed on the diencephalic brain sections. The immunoreactive somata were investigated for morphometry and counted using the optical fractionator method, throughout the confocal microscopy. Qualitative and quantitative research confirmed two separate populations of immunoreactive neurons. The modifications detected in the preoptic nucleus revealed that somata were more numerous, significantly smaller in size, and more excitable during the maturing phase but decreased, becoming slightly bigger and less excitable in the egg-laying stage. This may indicate that the catecholaminergic preoptic nucleus is involved in the control of reproduction, regulating both the onset of puberty and the imminent spawning. In contrast, somata in the suprachiasmatic nucleus grew in size and underwent turnover in morphometry, increasing the total number from the immature-virgin to maturing stage, with similar values in the more advanced maturity stages. These changes were not linked to a reproductive role. These findings provide new valuable information on Chondrichthyes, suggesting the existence of an additional brain system implicated in the integration of internal and environmental cues for reproduction.

Neural connections of the torus semicircularis in the adult Zebrafish.

The torus semicircularis (TS) of teleosts is a key midbrain center of the lateral line and acoustic sensory systems. To characterize the TS in adult zebrafish, we studied their connections using the carbocyanine tracers applied to the TS and to other related nuclei and tracts. Two main TS nuclei, central and ventrolateral, were differentiable by their afferent connections. From central TS, (TSc) numerous toropetal cells were labeled bilaterally in several primary octaval nuclei (anterior, magnocellular, descending, and posterior octaval nuclei), in the secondary octaval nucleus, in the caudal octavolateralis nucleus, and in the perilemniscular region. In the midbrain, numerous toropetal cells were labeled in the contralateral TSc. In the diencephalon, toropetal cells labeled from the TSc were observed ipsilaterally in the medial prethalamic nucleus and the periventricular posterior tubercle nucleus. TSc toropetal neurons were also labeled bilaterally in the hypothalamic anterior tuberal nucleus (ATN) and ipsilaterally in the parvicellular preoptic nucleus but not in the telencephalon. Tracer application to the medial octavolateralis nucleus revealed contralateral projections to the ventrolateral TS (TSvl), whereas tracer application to the secondary octaval nucleus labeled fibers bilaterally in TSc and neurons in rostral TSc. The TSc sends ascending fibers to the ipsilateral lateral preglomerular region that, in turn, projects to the pallium. Application of DiI to the optic tectum labeled cells and fibers in the TSvl, whereas application of DiI to the ATN labeled cells and fibers in the TSc. These results reveal that the TSvl and TSc are mainly related with the mechanosensory lateral line and acoustic centers, respectively, and that they show different higher order connections.

Acute and Reversible Hypothalamic Symptoms in a Lateral Head Impact Mouse Model of Mild Traumatic Brain Injury.

Central autonomic and endocrine dysfunctions following traumatic brain injury (TBI) are believed to involve the hypothalamus; however, underlying mechanisms are unknown. Although chronic deficits might be caused by irreversible tissue damage, various neuroendocrine and autonomic symptoms are only observed transiently, suggesting they might result from a temporary alteration in the activity of hypothalamic neurons. We therefore examined if a mouse model of mild TBI could induce reversible autonomic phenotypes and cause acute changes in c-Fos expression within corresponding regions of the hypothalamus. Adult C57Bl/6 male mice were lightly anesthetized with isoflurane and subjected to TBI by lateral head impact using a Gothenburg impactor. Mice treated the same way, but without the head impact served as controls (shams). We monitored body weight and core body temperature by infrared thermography and performed immunohistochemistry against c-Fos in various regions of the hypothalamus. We determined that a projectile velocity of 9 m/s significantly delayed recovery from the anesthesia without inducing skull fractures and signs of discomfort disappeared within 3 h, as assessed by grimace scale. Compared with shams, TBI mice displayed a rapid decrease in core body temperature which resolved within 48 h. Daily body weight gain was also significantly lower in TBI mice on the day following injury but recovered thereafter. c-Fos analysis revealed a significantly higher density of c-Fos-positive cells in the paraventricular nucleus and a significantly lower density in the median preoptic nucleus and medial preoptic area. We conclude that mild TBI induced by a single lateral head impact in mice at 9 m/s produces acute and reversible symptoms associated with hypothalamic dysfunction accompanied by significant changes in c-Fos expression within relevant areas of the hypothalamus. These findings support the hypothesis that a temporary alteration of neuronal activity may underlie the expression of reversible central autonomic and neuroendocrine symptoms.