Localization and neuronal response of RFamide related peptides in the rat central nervous system

RFamide related peptides (RFRP)-1 and RFRP-3 are neuropeptides derived from the same preproprotein. We have examined the distribution of RFRP-1 and RFRP-3 immunoreactivities (irs) in the rat central nervous system using specific antibodies. Neuronal cell bodies containing both RFRP-1 and RFRP-3 were detected within the caudal portion of the hypothalamus, the periventricular nucleus (PerVN), and the portion around or above the ventromedial nucleus of the hypothalamus. Both immunohistochemical and in situ hybridization analyses showed that neurons containing RFRP immunoreactivity and mRNA were distinct from those of neuropeptide FF, which contains the same structure at the C-terminus, Pro-Glu-Arg-Phe-NH2, as RFRP-3. Fibers containing both RFRP-1 and RFRP-3 were widely distributed in the brain: the lateral septal nucleus in the telencephalon, the paraventricular thalamic nucleus, various hypothalamic nuclei, the periaqueductal gray in the midbrain, the parabrachial nucleus in the pons, and the nucleus tractus solitarius (NTS) in the medulla oblongata. Only RFRP-1-ir was detected within the posterior gray horn in the spinal cord. Only RFRP-3-ir was detected in several thalamic nuclei and the spinal cord, especially at the posterior intermediate sulcus and within the anterior gray horn. Intracerebroventricular administration of RFRPs induced c-Fos expression in the anterior portion of the NTS, locus coeruleus, the nucleus of incertus, supraoptic nucleus, PerVN and the arcuate nucleus of the hypothalamus. These results show that RFRP-1 and RFRP-3 are widely distributed in the rat central nervous system and might be involved in various functions such as the neuroendocrine system or pain modulation.     

Sex differences in the regulation of progesterone receptor isoforms expression in the rat brain

We studied the effects of estradiol (E2) and progesterone (P) on progesterone receptor (PR) isoforms gene expression in the brain of ovariectomised female and gonadectomised male rats by RT-PCR analysis. In female rats, the expression of both PR isoforms was induced by E2 and down-regulated by P in the hypothalamus, whereas in the preoptic area these changes were only observed in PR-B isoform. On the contrary, in the hippocampus and the olfactory bulb, PR-A was the isoform induced by E2. In these regions, P did not modify the expression of any PR isoform. In the cerebellum and the frontal cortex of female rats, no treatment modified PR isoforms expression. In contrast with female rats, in the male rat brain, PR isoforms expression was only modified in the cerebellum, where PR-A was induced by E2. These results demonstrate a clear sexual dimorphism in the regulation of PR isoforms expression by sex steroid hormones in the rat brain, suggesting that this sex difference contributes to the sexually dimorphic effects of P in the rat brain.     

Immunocytochemical localization of substance P receptor in hypothalamic oxytocin-containing neurons of C57 mice

With the use of double immunofluorescence, we have examined the distribution of oxytocin-containing neurons that express substance P receptor (SPR) in the hypothalamus of C57 mice. The distribution of oxytocin-like immunoreactive neurons overlapped with that of SPR-like immunoreactive neurons in the paraventricular nucleus and supraoptic nucleus of the hypothalamus. Neurons showing both oxytocin- and SPR-like immunoreactivities were predominantly found in both nuclei. A few neurons that were double-labeled with oxytocin- and SPR-like immunoreactivities were also scattered in the hypothalamic periventricular and preoptic regions. Semi-quantitative analysis indicated that about 94% of the oxytocin-like neurons displayed SPR-like immunoreactivity. These double-labeled cells constituted about 91% of the SPR-like neurons in the aforementioned regions. The present study provides morphological evidence for tachykinin-induced modulation of oxytocin-containing neurons as mediated by substance P receptor in the hypothalamus of mammals.     

2,3,7,8-Tetrachlorodibenzo-p-dioxin treatment induces c-Fos expression in the forebrain of the Long-Evans rat

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is one of the most toxic environmental pollutants. In the present study, we examined c-Fos expression in the central nervous system (CNS) after administration of a lethal dose of TCDD to the adult Long-Evans rat to clarify if the CNS participates in TCDD-induced intoxication. A single dose of TCDD (dissolved in olive oil, 50 microg/kg) or olive oil alone was administered to the rats by gavage. Animals were allowed to survive for 1 day to 5 weeks. Three days after the administration, a significantly large number of Fos-immunopositive cells were found in the hypothalamus (i.e. dorsomedial hypothalamic nucleus, paraventricular hypothalamic nucleus, medial preoptic nucleus), central amygdaloid nucleus and bed nucleus of the stria terminalis. These results suggest that some TCDD toxicity may be induced by its direct action on the CNS.     

Long-term effects of St. John's wort and hypericin on monoamine levels in rat hypothalamus and hippocampus

Hypericum perforatum L. (St. John’s wort) is one of the leading psychotherapeutic phytomedicines and, because of this, great effort has been devoted to clarifying its mechanism of action. Chronic effects of St. John’s wort and hypericin, one of its major active compounds, on regional brain amine metabolism have not been reported yet. We used a high-performance liquid chromatography system to examine the effects of short-term (2 weeks) and long-term (8 weeks) administration of imipramine, Hypericum extract or hypericin on regional levels of serotonin (5-HT), norepinephrine, dopamine and their metabolites in the rat brain. We focused our interest on the hypothalamus and hippocampus, as these brain regions are thought to be involved in antidepressant drug action. Imipramine (15 mg/kg, p.o.), Hypericum extract (500 mg/kg, p.o.), and hypericin (0.2 mg/kg, p.o.) given daily for 8 weeks significantly increased 5-HT levels in the hypothalamus (P<0.05). The 5-HT turnover was significantly lowered in both brain regions after 8 weeks of daily treatment with the Hypericum extract (both P<0.05). Consistent changes in catecholamine levels were only detected in hypothalamic tissues after long-term treatment. Comparable to imipramine, Hypericum extract as well as hypericin significantly decreased 3,4-dihydroxyphenylacetic acid and homovanillic acid levels in the hypothalamus (P<0.01). Our data clearly show that long-term, but not short-term administration of St. John’s wort and its active constituent hypericin modify levels of neurotransmitters in brain regions involved in the pathophysiology of depression.     

Role of preoptic second messenger systems (cAMP and cGMP) in the febrile response

The present study aimed to test the hypothesis that a decrease in preoptic cAMP mediates fever. To this end, body core temperature (T(c)) of unanesthetized, freely moving rats was monitored by biotelemetry before and after pharmacological modulation of the cAMP pathway, and cAMP levels in the anteroventral third ventricular region (AV3V), where the preoptic region (POA) is located, were determined. We observed that intra-POA administration of the cAMP agonist dibutyryl-cAMP (Db-cAMP, 40 microg) reduced T(c). PGE(2) (the proximal mediator of fever, 200 ng) raised T(c) with a concomitant decrease in AV3V cAMP levels from 22.7+/-1.8 to 17.0+/-1.0 fmol/microg protein. Moreover, PGE(2)-induced fever was impaired by the phosphodiesterase inhibitor aminophylline. In order to verify the interaction between the cAMP- and cGMP-dependent pathways in the POA, we then co-injected Db-cAMP and 8-Br-cGMP into the POA. As a result, 8-Br-cGMP augmented the drop in T(c) evoked by Db-cAMP. Lastly, we observed that intra-POA co-microinjection of the protein kinase A inhibitor (Rp-cAMPS, 1 microg) with the protein kinase G inhibitor (Rp-cGMPS, 1 microg), mimicking the effects of reduced production of cAMP and cGMP, respectively, produced a fever-like response. In summary, the present data support that a decrease in the levels of cAMP and cGMP in the POA is associated with the genesis of fever.     

Hypothalamic c-fos-like immunoreactivity in high-fat diet-induced obese and resistant mice

Some C57Bl/6 mice become obese, whereas others remain lean when raised on a high-fat diet. The mechanisms underlying this interindividual susceptibility to diet-induced obesity remain unknown. Because hypothalamus plays a major role in the regulation of body weight, this study was conducted to identify the differences of hypothalamic neuronal activity between diet-induced obese and diet-resistant mice. Using c-fos as a marker, this study showed that diet-induced obese mice significantly increased c-fos-like immunoreactive neurons in the dorsal part of lateral hypothalamus (+183%) and dorsomedial hypothalamic nucleus (+87.5%) compared with diet-resistant mice. Furthermore, switching from high fat to low fat, or high n-3 polyunsaturated fatty acid diet, significantly decreased body weight gain (-35.7% and -31.0%), overall fat storage (-63.4% and -59.6%), and c-fos-like immunoreactive neurons in the dorsal part of lateral hypothalamus (-76.5% and -64.7%) and dorsomedial hypothalamic nucleus (-73.3% and -56.7%) in diet-induced obese mice, respectively. The present study also showed that the ratio of serum leptin/fat mass was threefold higher in the diet-resistant mice than in the diet-induced obese mice, which may be responsible for the less fat storage in the diet-resistant mice. The current data further confirm that the increased neuronal activity in the key autonomic regulatory centers may contribute to the excessive fat storage in diet-induced obese mice. Moreover, both high-fat diet-induced excessive fat storage and the altered hypothalamic neuronal activity may be largely corrected by reducing dietary fat content or replacing it with non-obesogenic fat.     

The effect on opioid peptides in the rat brain, after chronic treatment with the anabolic androgenic steroid, nandrolone decanoate

In recent years, an increase in abuse of anabolic androgenic steroids (AAS) has been seen among individuals not directly connected to sports. Clinical evidence suggests that abuse of these steroids may result in profound changes in personality, expressed by depressive symptoms, irritability and increased aggression. It is still unknown whether these alterations are related to changes in any particular transmitter system or whether they are persistent or reversible. In this study we focused on AAS effect on the endogenous dynorphin and enkephalin system in the brain. Male rats were given intramuscular injections of the AAS nandrolone decanoate (15 mg/kg), once daily for 2 weeks. The levels of the opioid peptide immunoreactivities (ir) were assessed by radioimmunoassay in two groups immediately after the treatment and in two other groups after additional 3 weeks without any drug treatment (recovery period). The result indicates that chronic AAS treatment increased the activity in the dynorphin B- and Met-enkephalin-Arg(6)Phe(7)-ir in the hypothalamus, striatum and periaqueductal gray (PAG) compared to controls. In addition, the steroid induced an imbalance between the dynorphin and the enkephalin opioid system in the nucleus accumbens, hypothalamus and PAG. This imbalance remained after the recovery period. Since increased peptide activity was found in brain regions regulating emotions, dependence, defensive reactions and aggression, it was suggested that the actual endogenous opioid systems are involved in previously reported AAS-induced changes in these behaviours.