This study was designed to determine the possible role of brain glucagon‐like peptide‐1 (GLP‐1) receptors in feeding behavior. In situ hybridization showed colocalization of the mRNAs for GLP‐1 receptors, glucokinase, and GLUT‐2 in the third ventricle wall and adjacent arcuate nucleus, median eminence, and supraoptic nucleus. These brain areas are considered to contain glucose‐sensitive neurons mediating feeding behavior. Because GLP‐1 receptors, GLUT‐2, and glucokinase are proteins involved in the multistep process of glucose sensing in pancreatic β cells, the colocalization of specific GLP‐1 receptors and glucose sensing‐related proteins in hypothalamic neurons supports a role of this peptide in the hypothalamic regulation of macronutrient and water intake. This hypothesis was confirmed by analyzing the effects of both systemic and central administration of GLP‐1 receptor ligands. Acute or subchronic intraperitoneal administration of GLP‐1 (7–36) amide did not modify food and water intake, although a dose‐dependent loss of body weight gain was observed 24 h after acute administration of the higher dose of the peptide. By contrast, the intracerebroventricular (i.c.v.) administration of GLP‐1 (7–36) amide produced a biphasic effect on food intake characterized by an increase in the amount of food intake after acute i.c.v. delivery of 100 ng of the peptide. There was a marked reduction of food ingestion with the 1,000 and 2,000 ng doses of the peptide, which also produced a significant decrease of water intake. These effects seemed to be specific because i.c.v. administration of GLP‐1 (1–37), a peptide with lower biological activity than GLP‐1 (7–36) amide, did not change feeding behavior in food‐deprived animals. Exendin‐4, when given by i.c.v. administration in a broad range of doses (0.2, 1, 5, 25, 100, and 500 ng), proved to be a potent agonist of GLP‐1 (7–36) amide. It decreased, in a dose‐dependent manner, both food and water intake, starting at the dose of 25 ng per injection. Pretreatment with an i.c.v. dose of a GLP‐1 receptor antagonist [exendin (9–39); 2,500 ng] reversed the inhibitory effects of GLP‐1 (7–36) amide (1,000 ng dose) and exendin‐4 (25 ng dose) on food and water ingestion. These findings suggest that GLP‐1 (7–36) amide may modulate both food and drink intake in the rat through a central mechanism.

Evidence that glucagon‐like peptide‐1 (GLP‐1) (7–36) amide functions as a novel neuropeptide prompted us to study the gene expression of its receptor in rat brain. Northern blot analysis showed transcripts of similar size in RINm5F cells, hypothalamus, and brainstem. First‐strand cDNA was prepared by using RNA from hypothalamus, brainstem, and RINm5F cells and subsequently amplified by PCR. Southern blot analysis of the PCR products showed a major 1.4‐kb band in all these preparations. PCR products amplified from hypothalamus were cloned, and the nucleotide sequence of one strand was identical to that described in rat pancreatic islets. In situ hybridization studies showed specific labeling in both neurons and glia of the thalamus, hypothalamus, hippocampus, primary olfatory cortex, choroid plexus, and pituitary gland. In the hypothalamus, ventromedial nuclei cells were highly labeled. These findings indicate that GLP‐1 receptors are actually synthesized in rat brain. In addition, the colocalization of GLP‐1 receptors, glucokinase, and GLUT‐2 in the same areas supports the idea that these cells play an important role in glucose sensing in the brain.

The influence of gonadal steroids on insulin-like growth factor I (IGF-I)-like immunoreactivity was assessed in the rat arcuate nucleus, an area of the hypothalamus that regulates pituitary secretion. IGF-I-like immunoreactivity was observed in hypothalamic cells with the morphological aspects of tanycytes and astrocytes. The surface density of IGF-I-like immunoreactive glia increased with puberty in the arcuate nucleus of male and female rats, while decreasing with age in other brain areas. Gender differences in the surface density of IGF-I-like immunoreactive glia were detected in adult animals, with males and androgenized females having significantly higher values than normal females. In the latter, the surface density of IGF-I-like immunoreactive glia was increased in the afternoon of proestrus and in the morning of estrus compared to the morning of proestrus, diestrus and metestrus. In addition, IGF-I-like immunoreactivity showed a dose-dependent increase in ovariectomized rats injected with 17β-estradiol, but not in those receiving 17α-estradiol. The effect of 17β-estradiol was blocked by simultaneous administration of progesterone, while this hormone alone had no effect. These results indicate that IGF-I-like immunoreactivity in arcuate glial cells is affected by the hormonal environment and suggest that IGF-I-like immunoreactive glia may be involved in neuroendocrine events within the hypothalamus.

The secretory pattern of GH secretion is markedly sexually dimorphic in the adult rat. The patterning of GH secretion is determined by the coordinated activity of somatostatin (SS)- and GH-releasing hormone (GHRH)-containing neurosecretory cells located in the hypothalamus. In this study we examined whether there is sexual dimorphism in the expression of the SS and GHRH genes and, if so, at what developmental stage this becomes evident. To address these questions, we measured SS messenger RNA (mRNA) levels in neurons of the periventricular nucleus and GHRH mRNA levels in the arcuate nucleus and ventromedial nucleus of the hypothalamus in male and female rats at 10, 25, 35, and 75 days of age. Using in situ hybridization and a computerized image analysis system, we measured SS mRNA and GHRH mRNA signal levels in individual neurons and compared these levels among the different age groups. We found that male animals had significantly higher levels of SS mRNA than females at every age. Similarly, males had higher GHRH mRNA levels than females; however, this difference was statistically significant only at 10 and 75 days of age. Developmental changes in GHRH mRNA levels were similar for both sexes, with GHRH message levels increasing gradually over the course of maturation. SS mRNA signal levels also changed over the course of development in both male and female animals. In the male rat, SS mRNA levels increased significantly between 10 and 25 days of age and declined significantly between 35 and 75 days of age. In the female rat, SS mRNA levels increased gradually between 10 and 35 days of age, then, as in the male, declined significantly between days 35 and 75. We conclude that sex differences and age-dependent changes in the expression of the SS and GHRH genes may subserve the sexual dimorphism and developmental alterations in the pattern of GH secretion in the rat.

Sex steroids play an important role in the development and functioning of the central nervous system (CNS); however, the mechanisms by which such hormones exert these effects are not well understood. We addressed the question as to whether sex steroids affect the development of the hypothalamus, at least in part, by acting as a trophic factor to modulate the number of neurons in the hypothalamus. To this end, primary hypothalamic cultures were prepared from the brains of embryonic (day 15) fetuses. Cultures received either 17β-es-tradiol (10^–12M) or vehicle 6 h after seeding and everyday throughout the study. As early as 24 h later, cultures receiving 17β-estradiol had significantly more neurons (44%, p < 0.001) than the control cultures. This effect not only continued throughout the duration of the study, but the difference between the two groups increased so that after 5 days, 17β-estradiol-treated cultures had 209% more neurons than control cultures (p < 0.001). Thus, addition of 17β-estradiol to fetal hypothalamic cultures produced a significant increase in the number of neurons surviving in vitro. The presence of glia was not required for this phenomenon, since the number of neurons surviving in glial-free cultures was also significantly increased by the addition of 17β-estradiol. The neuron survival promoting effect of 17β-estradiol was saturable and could be blocked by the estrogen antagonist tamoxifen (10^-7M). Testosterone (10^–10M), but not the nonaromatizable androgen dihydrotestosterone (10^–10M), could mimic the neuron survival-promoting effects of estradiol. Furthermore, estradiol had no significant effect on the in vitro survival of cerebral cortical neurons. These results suggest that one mechanism by which sex steroids may affect the development of the hypothalamus is through the modulation of the number of neurons that survive and that this effect is most likely mediated, at least in part, through the estrogen receptor.

No abstract

Recent studies have linked changes in peripheral chemokine concentrations to the presence of both addictive behaviors and psychiatric disorders. The present study further explore this link by analyzing the potential association of psychiatry comorbidity with alterations in the concentrations of circulating plasma chemokine in patients of both sexes diagnosed with alcohol use disorders (AUD). To this end, 85 abstinent subjects with AUD from an outpatient setting and 55 healthy subjects were evaluated for substance and mental disorders. Plasma samples were obtained to quantify chemokine concentrations [C–C motif (CC), C–X–C motif (CXC), and C–X3–C motif (CX3C) chemokines]. Abstinent AUD patients displayed a high prevalence of comorbid mental disorders (72%) and other substance use disorders (45%). Plasma concentrations of chemokines CXCL12/stromal cell-derived factor-1 (p < 0.001) and CX3CL1/fractalkine (p < 0.05) were lower in AUD patients compared to controls, whereas CCL11/eotaxin-1 concentrations were strongly decreased in female AUD patients (p < 0.001). In the alcohol group, CXCL8 concentrations were increased in patients with liver and pancreas diseases and there was a significant correlation to aspartate transaminase (r = +0.456, p < 0.001) and gamma-glutamyltransferase (r = +0.647, p < 0.001). Focusing on comorbid psychiatric disorders, we distinguish between patients with additional mental disorders (N = 61) and other substance use disorders (N = 38). Only CCL11 concentrations were found to be altered in AUD patients diagnosed with mental disorders (p < 0.01) with a strong main effect of sex. Thus, patients with mood disorders (N = 42) and/or anxiety (N = 16) had lower CCL11 concentrations than non-comorbid patients being more evident in women. The alcohol-induced alterations in circulating chemokines were also explored in preclinical models of alcohol use with male Wistar rats. Rats exposed to repeated ethanol (3 g/kg, gavage) had lower CXCL12 (p < 0.01) concentrations and higher CCL11 concentrations (p < 0.001) relative to vehicle-treated rats. Additionally, the increased CCL11 concentrations in rats exposed to ethanol were enhanced by the prior exposure to restraint stress (p < 0.01). Concordantly, acute ethanol exposure induced changes in CXCL12, CX3CL1, and CCL11 in the same direction to repeated exposure. These results clearly indicate a contribution of specific chemokines to the phenotype of AUD and a strong effect of sex, revealing a link of CCL11 to alcohol and anxiety/stress.

No abstract