Leptin: A Potential Link Between Obstructive Sleep Apnea and Obesity.

Chronic intermittent hypoxia (CIH), a pathophysiological manifestation of obstructive sleep apnea (OSA), is strongly correlated with obesity, as patients with the disease experience weight gain while exhibiting elevated plasma levels of leptin. This study was done to determine whether a relationship may exist between CIH and obesity, and body energy balance and leptin signaling during CIH. Sprague-Dawley rats were exposed to 96 days of CIH or normoxic control conditions, and were assessed for measures of body weight, food and water intake, and food conversion efficiency. At the completion of the study leptin sensitivity, locomotor activity, fat pad mass and plasma leptin levels were determined within each group. Additionally, the hypothalamic arcuate nucleus (ARC) was isolated and assessed for changes in the expression of proteins associated with leptin receptor signaling. CIH animals were found to have reduced locomotor activity and food conversion efficiency. Additionally, the CIH group had increased food and water intake over the study period and had a higher body weight compared to normoxic controls at the end of the study. Basal plasma concentrations of leptin were significantly elevated in CIH exposed animals. To test whether a resistance to leptin may have occurred in the CIH animals due to the elevated plasma levels of leptin, an acute exogenous (ip) leptin (0.04 mg/kg carrier-free recombinant rat leptin) injection was administered to the normoxic and CIH exposed animals. Leptin injections into the normoxic controls reduced their food intake, whereas CIH animals did not alter their food intake compared to vehicle injected CIH animals. Within ARC, CIH animals had reduced protein expression of the short form of the obese (leptin) receptor (isoform OBR100) and showed a trend toward an elevated protein expression of the long form of obese (leptin) receptor (OBRb). In addition, pro-opiomelanocortin (POMC) protein expression was reduced, but increased expression of the phosphorylated extracellular-signal-regulated kinase 1/2 (pERK1/2) and of the suppressor of cytokine signaling 3 (SOCS3) proteins was observed in the CIH group, with little change in phosphorylated signal transducer and activator of transcription 3 (pSTAT3). Taken together, these data suggest that long-term exposure to CIH, as seen in obstructive sleep apnea, may contribute to a state of leptin resistance promoting an increase in body weight.

Effect of estrogen on vagal afferent projections to the brainstem in the female.

The effects of 17ß-estradiol (E) on the distribution and density of brainstem projections of small or large diameter primary vagal afferents were investigated in Wistar rats using transganglionic transport of wheat germ agglutinin- (WGA; preferentially transported by non-myelinated afferent C-fibers; 2%), or cholera toxin B-subunit- (CTB, 5%; preferentially transported by large myelinated afferent A-fibers) conjugated horseradish peroxidase (HRP) in combination with the tetramethylbenzidine method in age matched ovariectomized (OVX) only or OVX and treated with E (OVX+E; 30 pg/ml plasma) females for 12 weeks. Additionally, these projections were compared to aged matched males. Unilateral microinjection of WGA-HRP into the nodose ganglion resulted in dense anterograde labeling bilaterally, with an ipsilateral predominance in several subnuclei of the nucleus of the solitary tract (NTS) and in area postrema that was greatest in OVX+E animals compared to OVX only and males. Moderately dense anterograde labeling was also observed in paratrigeminal nucleus (PAT) of the OVX+E animals. CTB-HRP produced less dense anterograde labeling in the NTS complex, but had a wider distribution within the brainstem including the area postrema, dorsal motor nucleus of the vagus, PAT, the nucleus ambiguus complex and ventrolateral medulla in all groups. The distribution of CTB-HRP anterograde labeling was densest in OVX+E, less dense in OVX only females and least dense in male rats. Little, if any, labeling was found within PAT in males using either WGA-or CTB-HRP. Taken together, these data suggest that small, non-myelinated (WGA-labeled) and large myelinated (CTB-labeled) diameter vagal afferents projecting to brainstem autonomic areas are differentially affected by circulating levels of estrogen. These effects of estrogen on connectivity may contribute to the sex differences observed in central autonomic mechanisms between gender, and in females with and without estrogen.

Hypothalamic orexin-A (hypocretin-1) neuronal projections to the vestibular complex and cerebellum in the rat.

Immunohistochemistry combined with retrograde tract-tracing techniques were used to investigate the distribution of orexin-A (OX-A)- and OX-A receptor-like (OX1) immunoreactivity within the vestibular complex and cerebellum, and the location of hypothalamic OX-A neurons sending axonal projections to these regions in the Wistar rat. OX-A immunoreactive fibers and presumptive terminals were found throughout the medial (MVe) and lateral (LVe) vestibular nuclei. Light fiber labeling was also observed in the spinal and superior vestibular nuclei. Within the cerebellum, dense fiber and presumptive terminal labeling was observed in the medial cerebellar nucleus (Med; fastigial nucleus), with less dense labeling in the interposed (Int) and lateral cerebellar nuclei (Lat; dentate nucleus). A few scattered OX-A immunoreactive fibers were also observed throughout the cortex of the paraflocculus. OX1-like immunoreactivity was found densely concentrated within LVe, moderate in MVe, and scattered within the spinal and superior vestibular nuclei. Within the cerebellum, OX1-like immunoreactivity was also observed densely within Med and in the dorsolateral aspects of Int. Additionally, OX1 like-labeling was found in Lat, and within the granular layer of the caudal paraflocculus cerebellar cortex. Fluorogold (FG) microinjected into these vestibular and cerebellar regions resulted in retrogradely labeled neurons throughout the ipsilateral hypothalamus. Retrogradely labeled neurons containing OX-A like immunoreactivity were observed dorsal and caudal to the anterior hypothalamic nucleus and extending laterally into the lateral hypothalamic area, with the largest number clustered around the dorsal aspects of the fornix in the perifornical area. A few FG OX-A like-immunoreactive neurons were also observed scattered throughout the dorsomedial, and posterior hypothalamic nuclei. These data indicate that axons from OX-A neurons terminate within the vestibular complex and deep cerebellar nuclei of the cerebellum and although the function of these pathways is unknown, they likely represent pathways by which hypothalamic OX-A containing neurons co-ordinate vestibulo-cerebellar motor and autonomic functions associated with ingestive behaviors.

Carotid chemoreceptor afferent projections to leptin receptor containing neurons in nucleus of the solitary tract.

Neurons expressing the leptin receptor (Ob-R) exist within the caudal nucleus of the solitary tract (NTS). Additionally, afferent neurons expressing the Ob-R have been identified within the nodose ganglion and NTS. Furthermore, systemic injections or focal injections of leptin directly into NTS potentiate the response of NTS neurons to carotid chemoreceptor activation. However, the distribution of carotid body afferents in relation to Ob-R containing neurons within NTS is not known. In this study, chemoreceptor afferent fibers were labeled following microinjection of the anterograde tract tracer biotinylated dextran amine (BDA) into the carotid body or petrosal/nodose ganglion of Wistar rats. After a survival period of 10-14 days, the NTS was processed for BDA and Ob-R immunoreactivity. Afferent axons originating in the carotid body were found to project to the lateral (Slt), gelantinosa (Sg), and medial (Sm) subnuclei of the NTS complex. A similar, but more robust distribution of BDA labeled fibers was observed in the NTS complex after injections into the petrosal/nodose ganglion. Carotid body BDA labeled fibers were observed in close apposition to Ob-R immunoreactive neurons in the region of Slt, Sg and Sm. In addition, a small number of carotid body afferents were found to contain both BDA and express Ob-R-like immunoreactivity within the regions of Slt, Sg and Sm. Taken together, these data suggest that leptin may modulate carotid chemoreceptor function not only through direct effects on NTS neurons, but also through a direct effect on carotid body primary afferent fibers that innervate NTS neurons.