CAT CEREBRAL ARTERIAL SMOOTH-MUSCLE CELLS EXPRESS CYTOCHROME-P450 4A2ENZYME AND PRODUCE THE VASOCONSTRICTOR 20-HETE WHICH ENHANCES L-TYPE CA2+ CURRENT

1. Cerebral arteries express cytochrome P450 4A enzymes (P450 4A) and produce 20-hydroxyeicosatetraenoic acid (20-HETE), a potent constricto r of pial arterioles. It is not known which cell type in the vessel ma ll is responsible for the formation of 20-HETE. We examined whether fr eshly isolated cerebral arterial muscle cells (VSMCs) express P450 4A and produce 20-HETE. We also studied the effect of 20-HETE on pressuri zed cerebral arteries and on whole-cell L-type Ca2+-current (I-Ca) rec orded in cat cerebral VSMCs. 2. Cat cerebral VSMCs incubated with [C-1 4]arachidonic acid ([C-14]AA) produced 20-HETE (3.9 +/- 1.1 pmol min(- 1) (mg protein)(-1)). 3. Reverse transcription-polymerase chain reacti on studies revealed that cat cerebral VSMCs express mRNA for P450 4A w hich metabolizes AA to 20-HETE. Cloning and sequencing of the cDNA amp lified from mRNA isolated from VSMCs showed > 96% amino acid homology to the rat and human P450 4A2 and 4A3. 4. 20-HETE (1-300 nM) induced a concentration-dependent constriction of cat cerebral arteries, which was inhibited by nifedipine. 5. Addition of 10 and 100 nM 20-HETE to t he bath increased peak I-Ca by 50 +/- 3 and 100 +/- 10%, respectively. This effect was not influenced by altering the frequency of depolariz ation. 20-HETE (100 nM) failed to increase I-Ca in the presence of nif edipine. 6. These results demonstrate that cat cerebral VSMCs express P450 4A enzyme, and produce 20-HETE which activates L-type Ca2+ channe l current to promote cerebral vasoconstriction.