Functional properties of Ca(v)1.3 (alpha(1D)) L-type Ca2+ channel splice variants expressed by rat brain and neuroendocrine GH(3) cells

Ca2+ enters pituitary and pancreatic neuroendocrine cells through dihydropy ridine-sensitive channels triggering hormone release. Inhibitory metabotrop ic receptors reduce Ca2+ entry through activation of pertussis toxin-sensit ive G proteins leading to activation of K+ channels and voltage-sensitive i nhibition of L-type channel activity. Despite the cloning and functional ex pression of several Ca2+ channels, those involved in regulating hormone rel ease remain unknown. Using reverse transcription-polymerase chain reaction we identified mRNAs encoding three alpha (1) (alpha (1A), alpha (1C), and a lpha (1D)), four beta, and one alpha (2)-delta subunit in rat pituitary GH( 3) cells; alpha (1B) and alpha (1S) transcripts were absent. GH(3) cells ex press multiple alternatively spliced alpha (1D) mRNAs. Many of the alpha (1 D) transcript variants encode "short" alpha (1D) (alpha (1D-S)) subunits, w hich have a QXXER amino acid sequence at their C termini, a motif found in all other al subunits that couple to opioid receptors. The other splice var iants identified terminate with a longer C terminus that lacks the QXXER mo tif (alpha (1D-L)). We cloned and expressed the predominant alpha (1D-S) tr anscript variants in rat brain and GH(3) cells and their alpha (1D-L) count erpart in GH(3) cells. Unlike alpha (1A) channels, alpha (1D) channels exhi bited current-voltage relationships similar to those of native GH(3) cell C a2+ channels, but lacked voltage-dependent G protein coupling. Our data dem onstrate that alternatively spliced alpha (1D) transcripts form functional Ca2+ channels that exhibit voltage-dependent, G protein-independent facilit ation. Furthermore, the QXXER motif, located on the C terminus of alpha (1D -S) subunit, is not sufficient to confer sensitivity to inhibitory G protei ns.