Evidence for 5-HT1D beta but not 5-HT1D alpha receptor subtype expression in canine large coronary arteries and saphenous vein

Abstract

Objective: 5-Hydroxytryptamine1D (5-HT1D) receptors are believed to play a major role in the vasoconstriction of vascular smooth muscle in human coronary arteries. However, unequivocal evidence as to which subtype of this receptor (5-HT1D alpha or 5-HT1D beta) is involved in these vasoconstrictory effects is lacking. The aim of this study was to identify in the dog the 5-HT1D receptor subtype encoding mRNAs expressed in several large coronary arteries and in the saphenous vein.

Methods: Degenerate oligonucleotide primers that selectively recognized only mammalian 5-HT1D alpha and 5-HT1D beta receptor sequences were used in RT-PCR experiments to study 5-HT1D receptor subtype expression in endothelium-denuded saphenous vein and large coronary arteries from beagle and alsatian dogs. Resulting PCR products were analysed and identified by Southern blots and sequencing.

Results: An identical PCR product whose sequence closely resembles that of the human 5-HT1D beta receptor (98% amino acid identity) was obtained from reverse-transcribed RNA isolated from either saphenous vein or coronary arteries, irrespective of dog race. Absence of 5-HT1D alpha expression was confirmed by Southern blot analysis. Control experiments using canine genomic DNA as template illustrated, nonetheless, that the primers chosen could amplify both 5-HT1D alpha and 5-HT1D beta sequences.

Conclusion: Using RT-PCR, we isolated from dog vascular smooth muscle a cDNA fragment whose nucleotide sequence would encode a previously-unreported canine homologue of the 5-HT1D beta receptor. We illustrated that this subtype is the only 5-HT1D receptor subtype expressed in dog saphenous vein and large coronary arteries. The implications of these findings are discussed in light of results from functional studies of 5-HT1-like receptor-mediated effects in these canine blood vessels.