AA (arachidonic acid) hydroxylation to 20-HETE (20-hydroxyeicosatetraenoic acid) influences renal vascular and tubular function. To identify the CYP (cytochrome P450) isoforms catalysing this reaction in the mouse kidney, we analysed the substrate specificity of Cyp4a10, 4a12a, 4a12b and 4a14 and determined sex- and strain-specific expressions. All recombinant enzymes showed high lauric acid hydroxylase activities. Cyp4a12a and Cyp4a12b efficiently hydroxylated AA to 20-HETE with V(max) values of approx. 10 nmol x nmol(-1) x min(-1) and K(m) values of 20-40 microM. 20-Carboxyeicosatetraenoic acid occurred as a secondary metabolite. AA hydroxylase activities were approx. 25-75-fold lower with Cyp4a10 and not detectable with Cyp4a14. Cyp4a12a and Cyp4a12b also efficiently converted EPA (eicosapentaenoic acid) into 19/20-OH- and 17,18-epoxy-EPA. In male mice, renal microsomal AA hydroxylase activities ranged between approx. 100 (NMRI), 45-55 (FVB/N, 129 Sv/J and Balb/c) and 25 pmol x min(-1) x mg(-1) (C57BL/6). The activities correlated with differences in Cyp4a12a protein and mRNA levels. Treatment with 5alpha-dihydrotestosterone induced both 20-HETE production and Cyp4a12a expression more than 4-fold in male C57BL/6 mice. All female mice showed low AA hydroxylase activities (15-25 pmol x min(-1) x mg(-1)) and very low Cyp4a12a mRNA and protein levels, but high Cyp4a10 and Cyp4a14 expression. Renal Cyp4a12b mRNA expression was almost undetectable in both sexes of all strains. Thus Cyp4a12a is the predominant 20-HETE synthase in the mouse kidney. Cyp4a12a expression determines the sex- and strain-specific differences in 20-HETE generation and may explain sex and strain differences in the susceptibility to hypertension and target organ damage.

Wall stretch is a major stimulus for the myogenic response of small arteries to pressure. Recent findings suggest that G protein‐coupled receptors can elicit a stretch response. Our aim was to determine if angiotensin II type 1 receptors (AT1R) in vascular smooth muscle cells (VSMC) exert mechanosensitivity and identify the downstream ion channel mediators of myogenic vasoconstriction. We used mice deficient in AT1R signaling molecules and putative ion channel targets, namely AT1R, angiotensinogen, TRPC6 channels or subtypes of the KCNQ (Kv7) gene family (KCNQ3, 4 or 5). We identified a mechano‐sensing mechanism in mesenteric arteries and the renal circulation that relies on coupling of the AT1R subtype a (AT1aR) to a Gq/11‐protein as a critical event to accomplish the myogenic response. The mechano‐activation occurs after block of AT1R, and in the absence of angiotensinogen or TRPC6. Activation of AT1aR suppresses XE991‐sensitive Kv channel currents in VSMCs, blocking these channels enhances mesenteric and renal myogenic tone. Although KCNQ3, 4 and 5 are expressed in VSMCs, XE991‐sensitive K+ current and myogenic contractions persist in arteries deficient in these channels. Our results provide evidence that myogenic responses of mouse mesenteric and renal arteries rely on ligand‐independent mechano‐activation of AT1aR. This signal relies on an ion channel distinct from TRPC6 or KCNQ3, 4 or 5. Grant Funding Source: Supported by Deutsche Forschungsgemeinschaft (DFG)

Objective-Epoxyeicosatrienoic acids (EETs) serve as endothelial-derived hyperpolarizing factors (EDHF), but may also affect vascular function by other mechanisms. We identified a novel interaction between EETs and endothelial NO release using soluble epoxide hydrolase (sEH) Ϫ/Ϫ and ϩ/ϩ mice. Methods and Results-EDHF responses to acetylcholine in pressurized isolated mesenteric arteries were neither affected by the sEH inhibitor, N-adamantyl-NЈ-dodecylurea (ADU), nor by sEH gene deletion. However, the EDHF responses were abolished by catalase and by apamin/charybdotoxin (ChTx), but not by iberiotoxin, nor by the cytochrome P450 inhibitor PPOH. All four EETs (order of potency: 8,9-EET Ͼ14,15-EETϷ5,6-EET Ͼ11,12-EET) and all 4 dihydroxy derivatives (14,15-DHETϷ8,9-DHETϷ11,12-DHET Ͼ5,6-DHET) produced dose-dependent vasodilation. Endothelial removal or L-NAME blocked 8,9-EET and 14,15-DHET-dependent dilations. The effects of apamin/ChTx were minimal. 8,9-EET and 14,15-DHET induced NO production in endothelial cells. ADU (100 g/mL in drinking water) lowered blood pressure in angiotensin II-infused hypertension, but not in L-NAME-induced hypertension. Blood pressure and EDHF responses were similar in L-NAME-treated sEH ϩ/ϩ and Ϫ/Ϫ mice. Conclusions-Our data indicate that the EDHF response in mice is caused by hydrogen peroxide, but not by P450 eicosanoids. Moreover, P450 eicosanoids are vasodilatory, largely through their ability to activate endothelial NO synthase (eNOS) and NO release. Key Words: eicosanoids Ⅲ soluble epoxide hydrolase Ⅲ NO synthase Ⅲ L-NAME Ⅲ EDRF T he endothelium releases nitric oxide (NO), prostacyclin, and endothelium-derived hyperpolarizing factor (EDHF). 1,2 Epoxyeicosatrienoic acids (EETs) are cytochrome P450 epoxygenase (CYP)-derived metabolites of arachidonic acid (AA) that may be EDHFs. 3,4 Other candidates include K ϩ ions and hydrogen peroxide (H 2 O 2 ). [5][6][7] Endothelial cell hyperpolarization spreads to adjacent vascular smooth muscle cells (VSMCs) through myo-endothelial gap junctions. 8,9 Calcium-activated potassium channels, most probably the SK4 (IK Ca ) and SK3 (SK Ca ) expressed on the endothelium, are the end-cellular gateway mediating hyperpolarization, and subsequent EDHF relaxation. 2,4,10 -13 EETs convincingly cause hyperpolarization. 14 -16 They can induce vasodilation in certain vascular beds by increasing the open-state probability of calcium-activated potassium (BK) channels. 4,15,17 The soluble epoxide hydrolase (sEH) metabolizes EETs to dihydroxy derivatives (DHET). sEH inhibition could enhance EET activity. 18 Blood pressure decreased in spontaneously hypertensive rats (SHR) given an sEH inhibitor. 19 sEH inhibition also lowered blood pressure in rats given angiotensin II (Ang II). 20 Thus, sEH could contribute to Ang IIinduced hypertension 21 and salt-sensitivity. 22 Even desoxycorticosterone acetate (DOCA)-salt hypertension was ameliorated with sEH inhibition. 23 Finally, male sEH genedeleted (Ϫ/Ϫ) mice had lower blood pressures than sEH ϩ/ϩ mice. 24 EETs coul...