Cardiovascular effects of intrahypothalamic injections of α-melanocyte stimulating hormone

Injection of α-melanocyte stimulating hormone (α-MSH, 0.6–12 nmol in 100–300 nl) into the rostral dorsomedial hypothalamic nucleus of the halothane anesthetized rat resulted in a 12% increase in heart rate (41 ± 4bpm) which was accompanied by a slight increase in blood pressure (5 ± 1mm Hg). The response was characterized by a gradual onset, with a peak increase at 7 ± 1 min and a duration of 51 ± 6 min. Tachyphylaxis to the response was apparent for at least 180 min following initial exposure to the peptide. In contrast to the increase in heart rate observed following α-MSH injection into the dorsomedial nucleus, injections into the medial preoptic, anterior, paraventricular or posterior hypothalamic nuclei had no significant effects on blood pressure and heart rate. These data suggest a possible role for brain α-MSH in the central control of heart rate at a site within the dorsomedial nucleus of the hypothalamus.     

Regional and subcellular localization of luteinizing hormone releasing hormone in the adult human brain

The localization of luteinizing hormone releasing hormone (LHRH) in the post mortem adult human brain was investigated. LHRH was highly concentrated in medial basal hypothalamic tissue (1.14 ng/mg protein); lower levels of LHRH were present in tissue from the optic chiasm (0.05 ng/mg protein) and mammillary bodies (0.07 ng/mg protein). The concentrations of LHRH in hypothalamic tissue of men and women were similar. LHRH was undetectable (< 0.001 ng/mg protein) in the frontal cerebral cortex and cerebellum. When homogenates of the medial basal hypothalamus were fractionated on continuous or discontinuous sucrose density gradients, LHRH was found to be associated with subcellular particles. Upon examination by transmission electron microscopy, we found that these subcellular particles resembled isolated neuron terminals, i.e., synaptosomes. Low to undetectable amounts of LHRH were found in the cytosol or the myelin + microsome fraction of the gradients. The results of these studies are supportive of the view that LHRH is highly concentrated in neuron terminals of the adult human brain and may, therefore, be a central neurotransmitter or neuromodulator in the human.     

Differential projections of two immunoreactive α-melanocyte stimulating hormone (α-MSH) neuronal systems in the rat brain

Immunocytochemical localization of α-MSH was performed in the brain of rats of which the arcuate nucleus has been destroyed by treatment with monosodium glutamate in the neonatal period. In these animals, α-MSH cell bodies normally found in the arcuate nucleus were almost completely absent. The reactive fibers found in the ACTH-β-LPH pathway were also markedly decreased. On the other hand, α-MSH cell bodies located in the dorsolateral hypothalamus as well as fibers located outside the ACTH-β-LPH pathway were not decreased. These results strongly suggest that α-MSH cell bodies in dorsolateral hypothalamus have projections completely different from those located in the arcuate nucleus.     

The distribution and projection of γ-melanocyte stimulating hormone in the rat brain: An immunohistochemical analysis

The distribution and projection of immunoreactive γ-melanocyte-stimulating hormone (γ-MSHI) in the rat brain was examined by indirect immunofluorescence using an antiserum against synthetic rat γ-MSH. The present study confirmed the presence of γ-MSHI neurons in the arcuate nucleus and further demonstrated that the n. communissuralis is a new γ-MSHI neurons-containing site. We also found a γ-MSHI fiber network in the hypothalamus, thalamus, amygdala, central gray matter of the midbrain and upper pons, and further demonstrated a much more extensive distribution of these fibers particularly in the medulla oblongata, an area previously thought not to contain γ-MSHI structures. The present observation on the normal distribution of γ-MSHI suggested the existence of two different systems: one is the arcuatofugal γ-MSH system and the other n. commissuralis γ-MSH system. Using experimental manipulations, we clearly established that γ-MSHI fibers in the forebrain, diencephalon, midbrain and upper pons originate from γ-MSHI neurons in the arcuate nucleus and those in the medulla oblongata from the n. commissuralis.     

The effect of α-melanocyte stimulating hormone (α-MSH) on the metabolism of blogenlc amines in the rat brain

The effect of the chronic administration of α-MSH on the incorporation of tritiated tyrosine into noradrenalin and dopamine and of tritiated tryptophan into serotonin was studied in different regions of the rat brain. α-MSH increased the incorporation of tritiated tryptophan into serotonin in the cortex and slightly decreased that of tyrosine into the dopamine in the hypothalamus. As the brain concentration of serotonin was unchanged in the animals treated with α-MSH, it is suggested that some of the changes in behavior, which other investigators have found following the administration of peptides containing the same sequence of amino acids to that found in MSH, could be associated with an increased turnover of cortical serotonin.     

Extrahypophyseal distribution of α-melanocyte stimulating hormone (α-MSH)-like immunoreactivity in postmortem brains from normal subjects and Alzheimer-type dementia patients

Using a sensitive double-antibody solid-phase enzyme immunoassay method α-melanocyte stimulating hormone-like immunoreactivity (α-MSH-LI) was measured in 21 regions of postmortem brains from 8 normal subjects and 5 patients with Alzheimer-type dementia (ATD). In the brains from the normal subjects, the highest concentration of α-MSH-LI was found in the hypothalamus. Relatively high concentration were also measured in the locus coeruleus, substantia innominata, substantia nigra, amygdala and medial nucleus of thalamus. α-MSH-LI in other regions was approximately1/100 of the hypothalamic content. This data is consistent with the existence of α-MSH in extrahypophyseal regions and indicates its regional distribution in the human brain. In the Alzheimer brains, although the temporal cortex and hippocampus had normal concentrations of α-MSH-LI, the cingulate cortex, caudate and substantia nigra showed significantly lower concentrations of α-MSH-LI than those of the control brains. This data suggests that further studies of α-MSH content in a larger number of ATD brains would be useful.     

Lateral hypothalamic innervation of the cerebral cortex: Immunoreactive staining for a peptide resembling but immunochemically distinct from pituitary/arcuate α-melanocyte stimulating hormone

The combination of retrograde transport of fluorescent dyes and indirect immunofluorescence has been used to study the putative neurotransmitter specificity of the tuberal lateral hypothalamic projection to the cerebral cortex. Injections of either fast blue or diamidino yellow dye into the cerebral cortex or hippocampus retrogradely labeled large, multipolar neurons scattered through the lateral hypothalamic area and zona incerta at the level of the ventromedial nucleus of the hypothalamus. Approximately 80% of these neurons stained immunohistochemically with an antiserum against α-melanocyte stimulating hormone (α-MSH). A second population of smaller, predominantly bipolar α-MSH-like immunoreactive neurons was seen in the arcuate nucleus and retrochiasmatic area, but none of these projected to the cerebral cortex. Immunohistochemical staining for ACTH (18–24), another proopiomelanocortin series peptide, or with an antiserum against α-MSH (4–10) demonstrated only the second of these cell groups. Our results indicate that the tuberal lateral hypothalamic projection to the cerebral cortex contains a substance similar but not identical to α-MSH, and that this material is probably not derived from the same proopiomelanocortin precursor as true α-MSH.     

Enhanced inhibitory feeding response to alpha-melanocyte stimulating hormone in the diet-induced obese rat

A dysregulation in the hypothalamic neuropeptide systems involved in the control of appetite has previously been shown in models of diet-induced obesity. In the present study, male Sprague-Dawley rats were rendered obese by a highly palatable cafeteria-style diet over 20 weeks, while control rats had access to standard laboratory chow. Feeding responses to alpha-melanocyte stimulating hormone (alpha-MSH), an anorectic peptide and neuropeptide Y (NPY), a potent orexigenic agent were investigated in diet-induced obese and control animals. In addition, endogenous hypothalamic peptide levels were determined in these animals. Intracerebroventricular injections of either 4 nmol alpha-MSH or saline vehicle were given 10 min prior to the onset of the dark phase. Diet-induced obese rats had significantly enhanced nocturnal inhibitory feeding responses to alpha-MSH (P<0.05). The orexigenic feeding response induced by 1 nmol NPY was similar for both groups. At sacrifice, both alpha-MSH and NPY peptide content were selectively reduced in the paraventricular nucleus (PVN) of these animals (P<0.05). Although diet-induced obesity had no effect on responses to NPY, the significantly greater inhibition of nocturnal feeding by alpha-MSH and reduction in PVN alpha-MSH peptide level, suggests melanocortinergic signalling may be reduced in obesity which may account for the hyperphagia of these animals when presented with a palatable diet.     

α-Melanocyte stimulating hormone discloses a stimulatory effect of β-endorphin on somatostatin release

The influence of alpha-melanocyte stimulating hormone (alpha-MSH) and beta-endorphin (beta-END) on the secretion of somatostatin (SRIF) from the median eminence (ME) was studied using an in vitro incubation system. The MEs from adult male rats were first preincubated at 37 degrees C for 30 min with constant shaking in 0.4 ml of Krebs-Ringer bicarbonate-glucose buffer (pH 7.4) containing bacitracin in an atmosphere of 95% O2/5% CO2. Medium was discarded and replaced by medium containing different doses of alpha-MSH, beta-END, or a fixed dose of alpha-MSH (10(-7) M or 10(-9) M) plus beta-END at various concentrations. By themselves alpha-MSH and beta-END did not alter basal SRIF release, but in the presence of alpha-MSH (10(-7) M) beta-END stimulated somatostatin release. This effect was significant at concentrations of beta-END of 10(-8) M and higher. The permissive effect of alpha-MSH was observed at a concentration as low as 10(-9) M, but in this case the stimulatory effect of beta-END became evident only at higher doses tested (10(-7) M). It is suggested that alpha-MSH and beta-END participate in the modulation of SRIF release. By themselves beta-END and alpha-MSH did not affect basal release of SRIF but in the presence of alpha-MSH, beta-END had a stimulatory effect on SRIF release. The mechanism for this interaction is unknown. The results are consistent with the possibility that beta-END neurons have stimulatory and inhibitory effects on SRIF release and that alpha-MSH, by blocking the inhibitory components, discloses the stimulatory effect of beta-END on SRIF release.     

Pro-opiocortin fragments in human and rat brain: β-endorphin and α-MSH are the predominant peptides

The ‘pro-opiocortin’ fragments, β-lipotropin, β-endorphin, ACTH and α-MSH, were estimated in discrete areas of rat and human brain and pituitaries by means of radioimmunoassay in combination with gelfiltration. These peptides exihibited parallel patterns of distribution, but with β-endorphin and α-MSH predominant in the brain of rat and man, and, in contrast, their respective precursors, β-LPH and ACTH predominant in the adenohypophysis of rat and man. These data may be indicative of important differences in post-translational processing of ‘pro-opiocortin’ between these contrasting tissues.     

Bilateral α-melanocyte stimulating hormonergic fiber system from zona incerta to cerebral cortex: combined retrograde axonal transport and immunohistochemical study

We employed a highly sensitive combination method of retrograde tracing and immunohistochemistry to identify an α-melanocyte-stimulating hormone (α-MSH)-containing fiber pathway from zona incerta to cerebral cortex. Biotin-horseradish peroxidase injected into the parietal cortex of the rat labeled a number of neurons in the zona incerta bilaterally, and stimultaneous staining with an α-MSH antiserum revealed that a part of these neurons are α-MSHergic.     

Immunocytochemically stained binding sites for oxytocin and α-melanocyte-stimulating hormone in rat brain following ventricular administration

Binding sites for oxytocin (OXT) and alpha-melanocyte-stimulating hormone (alpha-MSH) in brain of homozygous Brattleboro rats were immunocytochemically visualized after ventricular administration of the peptides by Accurel implants. Two patterns were found: 'ring type' staining in perineuronal structures was observed in CA1 and CA3 areas of ventral hippocampus and in subiculum for OXT implanted brains and a very weak staining in striatum for alpha-MSH-implanted brains; cytoplasmic staining of intracellular binding sites was observed in the bed nucleus of the stria terminalis (BST) in brains with OXT implants and in the anterodorsal thalamic nucleus (AD) and postcingulate cortex in brains with alpha-MSH implants. These localizations are different from those described for vasopressin binding sites in the same rat strain.     

Activation of supraoptic magnocellular neurons by gamma 2-melanocyte stimulating hormone (gamma 2-MSH)

The effects of intracarotid infusions of the peptide gamma₂-melanocyte stimulating hormone (γ₂-MSH) on electrophysiologically and immunohistochemically identified supraoptic nucleus (SON) units were investigated Over a wide dose range this agent always excited SON units, while control infusions of vehicle had no effect. Because neural responses invariably preceded blood pressure elevation, it appears that γ₂-MSH excitation of the magnocellular system was due to a direct effect on the central nervous system and was not a result of systemic cardiovascular responses. These results suggest a forebrain γ₂-MSH sensitive site in the activation of SON magnocellular neurons.     

α-Melanocyte-stimulating hormone structure-activity studies: Comparative analysis of melanotropic and CNS bioactivities

The structure-activity relationships in vitro of α-MSH (α-melanocyte-stimulating hormone, α-melanotropin) analogs as determined on normal and transformed (melanoma cell) melanocyte bioassays are summarized. Specifically, the characterization of potent and metabolically stable melanotropic agonist analogs and a newly discovered antagonist of α-MSH are highlighted. Comparison of these data versus the known structure-activity relationships of α-MSH related to CNS bioactivities suggests the existence of nonclassical α-MSH receptor-mediated pathways or, perhaps, a yet undefined endogenous neuropeptidergic pathway(s) having different selectivities for α-MSH analogs. In summary, several of the α-MSH analogs reported here may be useful molecular probes in future strategies aimed at the identification and systematic characterization of both peripheral and central α-MSH receptors.     

Plasma and cerebrospinal fluid α-MSH levels in the rat after hypophysectomy and stimulation of pituitary α-MSH secretion

Immunoreactive α-MSH was measured in cerebrospinal fluid (CSF) and plasma of rats. While treatment with haloperidol increased α-MSH levels in the plasma concentration of α-MSH in the CSF showed little change. Hypophysectomy also had little effect on the concentration of α-MSH in the CSF despite the fall in plasma α-MSH levels. This lack of correlation between α-MSH levels in the CSF and plasma suggests that the systemic circulation does not deliver α-MSH to the CSF. The apparently normal levels of α-MSH in the hypothalamus after hypophysectomy suggests that this tissue is able to synthesize α-MSH and it is possible that the hypothalamus is a source of the α-MSH in the CSF.     

Biological and immunological characterization of α-melanocyte-stimulating hormone (α-MSH) in two neuronal systems of the rat brain

Recent immunocytochemical studies have demonstrated the existence of two different neuronal systems containing α-MSH-like material in the brain: one originating from the arcuate nucleus and the other one from the dorsolateral hypothalamus. The aim of the present study was to further characterize α-MSH in these two discrete regions of the rat diencephalon. Intracerebroventricular administration of colchicine resulted in a marked decrease in the number of ACTH and β-endorphin nerve fibers and a significant reduction in ACTH and β-endorphin content in the dorsolateral hypothalamus. Conversely, colchicine treatment did not alter α-MSH, ACTH or β-endorphin content in the arcuate nucleus and did not significantly affect α-MSH concentration in the dorsal region. Reverse-phase high-performance liquid chromatography showed that the major α-MSH-like compound localized in the dorsal hypothalamus co-migrated exactly with synthetic α-MSH, whereas the arcuate nucleus contained 5 peptides cross-reacting with α-MSH antibodies, 4 of them being different from standard α-MSH. Significant amounts of biologically active melanotropin, which migrated on Sephadex G-25 columns like synthetic α-MSH, were also detected in both the arcuate nucleus and dorsolateral hypothalamus. Taken together, these data demonstrate that the α-MSH cell bodies located in the dorsolateral hypothalamus specifically produce authentic α-MSH, whereas the α-MSH cell bodies in the arcuate nucleus also contain ACTH, β-endorphin and several peptides immunologically related but not identical to α-MSH.