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Pituitary adenylate cyclase-activating peptide and vasoactive intestinal peptide receptor expression in immortalized
J Olcese1, C A McArdle, R Middendorff
1Institute for Hormone and Fertility Research, University of Hamburg, Germany.
The regulation of LHRH secretion is extraordinarily multifarious. To no small extent, this insight has been gained through studies using the immortalized hypothalamic LHRH neuronal line, GT1-7. In the present study, we examined these cells for potential expression of the receptors for the related peptides PACAP and VIP. By means of reverse transcription-polymerase chain reaction (RT-PCR) with PACAP receptor-specific primers, in combination with restriction enzyme analysis and cDNA sequencing, we were able to identify all PACAP-specific receptor splice variant forms with variable degrees of expression. Of the two nonselective VIP/PACAP receptors (i.e. VIP-R type I and II) only the latter isoform was detected by RT-PCR. In view of these results, we sought to establish whether PACAP and VIP receptors are functional in GT1-7 cells. Cyclic AMP (cAMP) accumulation after addition of PACAP-38 (or PACAP-27) was dose-dependent with maximal 3-fold increases. VIP also elevated cAMP with a similar potency. Phosphatidylinositol (PI) turnover was unaffected by either PACAP or VIP. Acute LHRH secretion was stimulated equally by nanomolar concentrations of both PACAP and VIP. These results point to PACAP and VIP having direct actions via the VIP2R on cAMP signalling and LHRH release, in addition to the known effects of these peptides on pituitary functions.
The regulation of LHRH secretion is extraordinarily multifarious. To no small extent, this insight has been gained through studies using the immortalized hypothalamic LHRH neuronal line, GT1-7. In the present study, we examined these cells for potential expression of the receptors for the related peptides PACAP and VIP. By means of reverse transcription-polymerase chain reaction (RT-PCR) with PACAP receptor-specific primers, in combination with restriction enzyme analysis and cDNA sequencing, we were able to identify all PACAP-specific receptor splice variant forms with variable degrees of expression. Of the two nonselective VIP/PACAP receptors (i.e. VIP-R type I and II) only the latter isoform was detected by RT-PCR. In view of these results, we sought to establish whether PACAP and VIP receptors are functional in GT1-7 cells. Cyclic AMP (cAMP) accumulation after addition of PACAP-38 (or PACAP-27) was dose-dependent with maximal 3-fold increases. VIP also elevated cAMP with a similar potency. Phosphatidylinositol (PI) turnover was unaffected by either PACAP or VIP. Acute LHRH secretion was stimulated equally by nanomolar concentrations of both PACAP and VIP. These results point to PACAP and VIP having direct actions via the VIP2R on cAMP signalling and LHRH release, in addition to the known effects of these peptides on pituitary functions.