Brain research. Molecular brain research最新文献

Prolonged exposure of rats to the synthetic cannabinoid receptor ligand, CP-55,940 (0.4 mg/kg, i.p. for 11 days), induced tolerance to analgesia, to the reduction in spontaneous locomotor activity and the incidence of splayed hind limbs. One hour after the last injection on day 11, the rats were killed and in situ hybridization was used to investigate the effect of treatment on G-protein alpha-subunit expression throughout the brain. Chronic cannabinoid exposure markedly reduced G alpha(s), G alpha(i) and G alpha(o) mRNA levels. The message for the alpha(s)-subunit was decreased in all the brain areas containing the basal autoradiographic signal; the decrease ranging from 25% in the thalamus to 45% in the mesencephalon. Also the basal G alpha(i) expression was reduced in tolerant rats showing the greatest decrease in the forebrain (63%) in the cerebellum (58%) and in the mesencephalon (38%). The reduction in G alpha(o) expression (25%) was more localized, being present only in the rostral portion of the brain (cortex, striatum and olfactory area). The alterations in alpha-subunits gene expression were not followed by any change in the amount of proteins. Our results indicate that, besides the receptor modification, alteration to the G-protein expression could be a molecular event associated with the development of cannabinoid tolerance.

The expression of mRNAs coding for tryptophan hydroxylase (TPOH), the first enzyme involved in melatonin synthesis, has been studied in the rat adult pineal gland at four different circadian time points by in-situ hybridization using an oligonucleotide probe. TPOH mRNAs were detected at all chosen time-points. The lowest level was seen at 13:00 h. Quantification of the hybridization signals demonstrated a significant increase in expression (+16.5%) between 13:00 h and 19:30 h, with the values at 23:30 h and 09:00 h being similar to that of 19:30 h. These results coincide with the nycthemeral rhythm of TPOH activity. Day/night rhythms in the rat pineal gland are controlled by noradrenaline release at lights-off, which results in an increase in cAMP levels and in gene activation. This study suggests that the TPOH gene might be one such gene whose activation is stimulated in this way.

The inhibitory glycine receptor (GlyR) is a pentameric receptor comprised of alpha and beta subunits, of which the beta subunit has not been characterised in humans. A 2106 bp cDNA, isolated from a human hippocampal cDNA library, contained an open reading frame of 497 amino acids which encodes the beta subunit of the human GlyR. The mature human GlyR beta polypeptide displays 99% amino acid identity with the rat GlyR beta subunit and 48% identity with the human GlyR alpha 1 subunit. Neither [3H]strychnine binding nor glycine-gated currents were detected when the human GlyR beta subunit cDNA was expressed in the human embryonic kidney 293 cell line. However, co-expression of the beta subunit cDNA with the alpha 1 subunit cDNA resulted in expression of functional GlyRs which showed a 4-fold reduction in the EC50 values when compared to alpha 1 homomeric GlyRs. Glycine-gated currents of alpha 1/beta GlyRs were 17-fold less sensitive than homomeric alpha 1 GlyRs to the antagonists picrotoxin, picrotoxinin and picrotin, providing clear evidence that heteromeric alpha 1/beta GlyRs were expressed. The beta subunit appears to play a structural rather than ligand binding role in GlyR function. Fluorescence in situ hybridisation was used to localise the gene encoding the human GlyR beta subunit (GLRB) to chromosome 4q32, a position syntenic with mouse chromosome 3. In situ hybridisation using the human GlyR beta subunit cDNA showed that the murine GlyR beta subunit gene (Glrb) maps to the spastic (spa) locus on mouse chromosome 3 at bands E3-F1. This is consistent with the recent finding that a mutation in the murine GlyR beta subunit causes the spa phenotype. It also raises the possibility that mutations in the human beta subunit gene may cause inherited disorders of the startle response.

The growth hormone (GH) secretory pattern is dependent on sex and developmental stage. It is generally accepted that in the male rat this pattern is markedly influenced by androgens secreted by the Leydig cells. Recent findings, however, point to the existence of other non-androgenic testicular factors produced by the Sertoli cells and which regulate in vivo the GH responses to growth hormone releasing hormone (GHRH). The aim of this work was to investigate the role played by non-androgenic testicular factors on hypothalamic somatostatin (SST) and GHRH mRNA levels. Seventy-day-old male Sprague-Dawley rats were used throughout the work. They were divided into five groups: (1) control rats; (2) gonadectomized rats; (3) gonadectomized rats supplemented with exogenous administration of dihydrotestosterone (DHT); (4) ethylene dimethane sulphonate (EDS)-treated rats; (5) EDS-treated rats supplemented with exogenous administration of DHT. EDS is a cytotoxic agent that specifically destroys the Leydig cells. The rats were killed after 15 days of treatment. Hypothalamic SST mRNA levels were determined by Northern blot and by in situ hybridization, and GHRH mRNA levels assessed by Northern blot. We found that selective removal of Leydig cells with EDS greatly reduced the SST mRNA content in the periventricular nucleus of the hypothalamus. These levels were significantly lower than those found in gonadectomized rats. Furthermore, replacement treatment with dihydrotesterone (DHT) did not completely restore SST mRNA levels in EDS-treated rats, contrasting with the complete recovery of SST mRNA levels in gonadectomized rats. On the other hand, gonadectomy and EDS treatment produced a significant reduction in GHRH mRNA levels. DHT administration reversed the action of gonadectomy, but did not restore GHRH mRNA content in EDS-treated rats. These data suggest that, in addition to testosterone, as yet unidentified non-androgenic testicular factors can significantly influence SST and GHRH mRNA levels. This may indicate that non-androgenic testicular factors acting at hypothalamic level may be important in the neuroregulation of GH secretion and in the maintenance of sexual dimorphism in GH secretory pattern.