RESUMEN
The histaminergic H3-receptor (H3R) controls histamine synthesis and release in the tuberomamillary nucleus. We evaluated the effects of stimulating or blocking of H(3)R on glutamate-decarboxylase 67 kDa (GAD-67) and galanin mRNA expression, two histamine co-transmitters.After in situ hybridization histochemistry (ISHH), we observed a colocalization of 100% between histidine decarboxylase (HDC) and GAD-67 or H3R and of 80 to 97% with galanin. Adult rats received an H3R agonist ((R)alpha-Methylhistamine) or antagonist (ciproxifan) and were sacrificed 1 or 3 hours later. Treatment effects on HDC, galanin and GAD-67 mRNA were studied by quantitative ISHH on serial sections. Treatment with the H3R agonist known to decrease histamine neuron activity initially reduced HDC and galanin gene expression but an inverse change, presumably reflecting a compensatory mechanism, was observed after 3 h on both markers. In contrast, the H3R antagonist known to activate histamine neurons, had opposite effects on the two markers, suggesting that co-transmitters are submitted to independent control mechanisms. Furthermore, GAD-67 mRNA levels were not significantly modified by these treatments.
Asunto(s)
Galanina/genética , Histamina/biosíntesis , Hipotálamo/efectos de los fármacos , Neuronas/efectos de los fármacos , Receptores Histamínicos H3/efectos de los fármacos , Ácido gamma-Aminobutírico/biosíntesis , Animales , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/fisiología , Glutamato Descarboxilasa/genética , Agonistas de los Receptores Histamínicos/farmacología , Antagonistas de los Receptores Histamínicos/farmacología , Histidina Descarboxilasa/genética , Área Hipotalámica Lateral/citología , Área Hipotalámica Lateral/efectos de los fármacos , Área Hipotalámica Lateral/metabolismo , Hipotálamo/citología , Hipotálamo/metabolismo , Imidazoles/farmacología , Isoenzimas/genética , Masculino , Metilhistaminas/farmacología , Neuronas/citología , Neuronas/metabolismo , ARN Mensajero/efectos de los fármacos , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Receptores Histamínicos H3/metabolismoRESUMEN
Membrane preparations of cells expressing the cloned rat hypothalamus melanocortin receptor, MC3, have been photoaffinity labelled using a radiolabelled photoreactive analogue of alpha-MSH, [125I-Tyr2,Nle4,D-Phe7,ATB-Lys11]alpha-MSH. SDS-PAGE followed by autoradiography showed a single band at 53-56 kDa for the native receptor or 35 kDa after deglycosylated with PNGase F, consistent with the predicted cDNA sequence. Receptor binding studies with alpha-MSH, gamma-MSH and [Nle4,D-Phe7]alpha-MSH established that alpha-MSH and gamma-MSH had similar affinities while [Nle4,D-Phe7]alpha-MSH bound 100 times more strongly. These results suggest that the receptor recognises the conserved 'core sequence' (-Met-Glu/Gly-His-Phe-Arg-Trp-) of MSH/ACTH peptides. The binding affinities of alanine-substituted analogues of alpha-MSH were determined to investigate the role of individual residues in ligand-receptor interactions. While in the terminal regions only the replacement of Tyr2 reduced the affinity of the peptide, replacement of Met4, Phe7, Arg8 and Trp9 within the peptide core led to a significant loss of affinity. Glu5 appeared unimportant for receptor recognition.