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The theory holds that the anterior pituitary in mammals receives humoral regulation. Previous studies have reported that the pars distalis of the anterior pituitary of several mammalian species contains substance P-, calcitonin gene-related peptide (CGRP)-, and galanin-like immunoreactive nerve fibers, but the origins of these nerve fibers are unclear. Removal of the pituitary gland, also called hypophysectomy, involves methods that access the pituitary gland via the transauricular or parapharyngeal pathways. However, these methods are not applicable for viral tracer injection to investigate the innervation of the anterior pituitary. The transauricular technique leads to inaccuracies in locating the pituitary gland, while the parapharyngeal approach causes high mortality in animals. Here, we introduce a protocol that accesses the pituitary gland in the rat via the transsphenoidal pathway. This method imitates surgical manipulations such as endotracheal intubation and sphenoid bone drilling, which involve the use of custom-made devices. Using the transsphenoidal pathway greatly improves the survival rate of rats because no additional dissection of blood vessels and nerves is required. Moreover, the pituitary gland can be viewed clearly and directly during the operation, making it possible to accurately inject pseudorabies virus (PRV) 152-expressing enhanced green fluorescent protein (EGFP) into the anterior or posterior pituitary, respectively. After injecting PRV 152 into the anterior pituitary, we found no evidence of direct innervation of the anterior pituitary in the rat brain. However, PRV 152 injection into the posterior pituitary revealed retrograde transneuronal cell bodies in many brain areas, including the CA1 field of the hippocampus, the basolateral amygdaloid nucleus, posterior part (BLP), the arcuate hypothalamic nucleus (Arc), the dorsal portion of the dorsomedial hypothalamic nucleus (DMD), the suprachiasmatic nucleus (SCh), and the subfornical organ (SFO). In the present study, we provide a description of a possible model of hypophysectomy or pituitary injection, and identify brain regions involved in regulating the rat pituitary gland using transneuronal retrograde cell body labeling with PRV., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be constructed as a potential conflict of interest., (Copyright © 2020 Li, Su, Zhao, Li, Xu, Gao, Sun, Yang, Jin and Ke.)

Orexin receptors type 1 (OX1R) and type 2 (OX2R) are G protein-coupled receptors whose structure is highly conserved in mammals. OX1R is selective for orexin A, and OX2R binds orexin A and orexin B with similar affinity. Orexin receptor expression was observed in human, rat, porcine, sheep as well as Xenopus laevis pituitaries, both in the adenohypophysis and in the neurohypophysis. The expression level is regulated by gonadal steroid hormones and GnRH. The majority of orexins reaching the pituitary originate from the lateral hypothalamus, but due to the presence of the receptors and the local production of orexins in the pituitary, orexins could deliver an auto/paracrine effect within the gland. Cumulative data indicate that orexins are involved in the regulation of LH, GH, PRL, ACTH, and TSH secretion by pituitary cells, pointing to orexins' effect on the functioning of the endocrine axes. Those hormones may also serve as a signal linking metabolic status with endocrine control of sleep, arousal, and reproduction processes., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Developing neurons are guided to their appropriate targets by specific guidance substances that have neurotrophic actions. The aim of the present study was to elucidate the mechanism by which hypothalamic neurons reach the pars intermedia (PI) by correlating the development of dopaminergic (DA) neurons arising in the periventricular nucleus (PeV) of fetal rats with the expression of brain-derived neurotrophic factor (BDNF) in the rat pituitary. The differentiation of DA neurons was observed by immunohistochemistry using an antibody against tyrosine hydroxylase (TH), whereas the ontogenesis of BDNF mRNA in the PI was examined by in situ hybridization and RT-PCR. Immunoreactive TH-neurons were first observed in the PeV at embryonic day (E) 16.5, following which time their axons elongated toward the pituitary. TH-positive reactions were observed in the connective tissue between the PI and the pars nervosa at E20.5. Innervation of the PI by TH-positive neurons was determined at postnatal day (P) 1.5; however, BDNF mRNA was first detected in the PI cells at E17.5, with an increase in its expression clearly visible at E21.5 and continuing high expression levels in the PI thereafter. These results suggest that BDNF is a specific guidance cue for DA neurons elongating from the PeV to the PI.

The neuroanatomical organization of neurons projecting to the pituitary and the origin of pituitary dopamine and nonapeptides were investigated in the brain of the Atlantic salmon (Salmo salar). Carbocyanine tract tracing in combination with tyrosine hydroxylase, arginine vasotocin and isotocin immunocytochemistry for double labelling revealed a previously unknown organization of hypophysiotrophic cell groups and their extrahypothalamic projections, and provide the first direct identification in a teleost fish of the origin of the dopaminergic and nonapeptidergic innervation of the pituitary. The present data include identification of (1) hypophysiotrophic neurons in the ventral telencephalon and in the periventricular preoptic nucleus, (2) large (magnocellular) vasotocinergic hypophysiotrophic neurons in the most rostral extension of the preoptic area, (3) a distinct neuronal group located in a supraoptic/suprachiasmatic position in the anterior periventricular nucleus, that seems to be the major source of dopaminergic innervation of the pituitary, (4) the nonapeptidergic hypophysiotrophic neurons in the preoptic nucleus, (5) hypophysiotrophic neurons in the ventral and posterior hypothalamus of which some are of liquor-contacting type, (6) projections from hypophysiotrophic and non-hypophysiotrophic neurons in the preoptic nucleus to extrahypothalamic areas such as thalamic and periventricular pretectal nuclei, and (7) subdivisions within the preoptic nucleus that exhibit different combinations of hypophysiotrophic and extrahypothalamic efferent connections. Together with previous studies of retinohypothalamic projections and neurochemical organization of hypothalamic/preoptic areas, the present data suggest that the preoptic nucleus and the anterior periventricular nucleus in teleosts possess functional subdivisions with features that resemble those of the paraventricular, supraoptic and suprachiasmatic nuclei of other vertebrates. In the Atlantic salmon, specific dopaminergic and nonapeptidergic neuronal subdivisions are proposed to play a role for photoperiod control of endocrine activity.

The activities of periventricular-hypophysial dopaminergic (DA) neurons were compared in male and female rats by measuring dopamine synthesis (accumulation of 3,4-dihydroxyphenylalanine [DOPA] after inhibition of L-aromatic amino acid decarboxylase) and metabolism (concentrations of 3,4-dihydroxyphenylacetic acid [DOPAC]) in terminals of these neurons in the intermediate lobe of the pituitary. For comparison, the synthesis and metabolism of dopamine in the neural lobe of the pituitary and median eminence were also determined. The concentrations of DOPAC and accumulation of DOPA were higher in females than in males in both the intermediate lobe and median eminence, revealing a sexual difference in the basal activity of periventricular-hypophysial and tuberoinfundibular DA neurons. In contrast, there were no differences between male and female rats in activity of DA neurons terminating in the neural lobe. One week following gonadectomy, DOPA accumulation in the median eminence was decreased in females and increased in males, but remained unchanged in the intermediate lobe. These results indicate that sexual differences in the activity of periventricular-hypophysial DA neurons terminating in the intermediate lobe are not dependent upon the presence of circulating gonadal steroids, and in this respect, these neurons differ from tuberoinfundibular DA neurons.

The effects of elevated circulating concentrations of prolactin were examined on neurochemical estimates of activity of dopaminergic neurons in the infundibulum/pituitary stalk of Holstein bull calves (8-10 weeks of age). Activity of these neurons was estimated by measuring the accumulation of dihydroxyphenylalanine, the immediate precursor of dopamine, 15 min after an intravenous injection of the aromatic L-amino acid decarboxylase inhibitor, 3-hydroxybenzylhydrazine. Subcutaneous injections of the dopamine antagonist haloperidol every 6 hr for 1 day increased serum concentrations of prolactin and accumulation of dihydroxyphenylalanine in the infundibulum/stalk. Intravenous infusions of prolactin for 1 or 9 days increased accumulation of dihydroxyphenylalanine in the infundibulum/stalk, indicating that these neurons remain responsive to elevated prolactin for at least 9 days. It is concluded that elevated concentrations of prolactin in blood stimulate dopaminergic neurons in the infundibulum/pituitary stalk of bull calves. We speculate that these neurons may be analogous to the tuberoinfundibular dopaminergic neurons that regulate prolactin in rats.

Using immunocytochemical methods at the electron microscope level, immunoreactivity for both melanin-concentrating hormone (MCH) and alpha-melanocyte-stimulating hormone (alpha-MSH) has been demonstrated in the carp neurohypophysis. A double-labelling technique, using colloidal gold probes of different sizes showed that immunoreactivity to both molecules coexists within the same neurosecretory granules in some neurones, while in other neurones the granules exhibit only MCH-like immunoreactivity. These observations suggest that the two immunoreactivities are attributable to separate molecules; if they are derived from the same precursor molecule, then this must be cleaved differently in the two sets of neurones. The absence of adrenocorticotropic hormone (ACTH)-like immunostaining in any neurosecretory granule might suggest the alpha-MSH-like molecule is not derived from the conventional pro-opiomelanocortin precursor.

5-Hydroxytryptamine (5-HT)-containing axons and terminals have been visualized in the neural and intermediate lobes of the rat pituitary gland, but the origin of these fibers remains in question. This study was designed to determine if 5-HT cell bodies in the brainstem or in the dorsomedial nucleus of the hypothalamus project to either of these pituitary lobes. Since lesions and electrical stimulation of 5-HT cell bodies decrease and increase, respectively, the rate of 5-HT synthesis in regions innervated by these cells, these techniques were employed. The in vivo rate of 5-HT synthesis was determined by quantifying the rate of accumulation of the 5-HT precursor, 5-hydroxytryptophan (5-HTP), in the neural and intermediate lobes of the pituitary gland 30 min after the administration of a decarboxylase inhibitor (NSD 1015, 100 mg/kg, i.p.). The application of 30 min of stimulating current (monophasic cathodal pulses of 1 ms duration and 0.3 mA current delivered at a frequency of 10 Hz) to electrodes implanted in the dorsal and median raphe nuclei increased the rate of 5-HT synthesis in both the neural and intermediate lobes of the pituitary gland. 5,7-Dihydroxytryptamine lesions of these nuclei altered neither 5-HTP accumulation nor 5-HT concentrations in the neural and intermediate lobes, but similar lesions of the nuclei raphe pontis and raphe magnus decreased both the concentration of 5-HT and the accumulation of 5-HTP in these pituitary regions.(ABSTRACT TRUNCATED AT 250 WORDS)