Objective-Ca2ϩ -influx through transient receptor potential (TRP) channels was proposed to be important in endothelial function, although the precise role of specific TRP channels is unknown. Here, we investigated the role of the putatively mechanosensitive TRPV4 channel in the mechanisms of endothelium-dependent vasodilatation. Methods and Results-Expression and function of TRPV4 was investigated in rat carotid artery endothelial cells (RCAECs) by using in situ patch-clamp techniques, single-cell RT-PCR, Ca 2ϩ measurements, and pressure myography in carotid artery (CA) and Arteria gracilis. In RCAECs in situ, TRPV4 currents were activated by the selective TRPV4 opener 4␣-phorbol-12,13-didecanoate (4␣PDD), arachidonic acid, moderate warmth, and mechanically by hypotonic cell swelling. Single-cell RT-PCR in endothelial cells demonstrated mRNA expression of TRPV4. In FURA-2 Ca 2ϩ measurements, 4␣PDD increased [Ca 2ϩ ] i by Ϸ140 nmol/L above basal levels. In pressure myograph experiments in CAs and A gracilis, 4␣PDD caused robust endothelium-dependent and strictly endothelium-dependent vasodilatations by Ϸ80% (K D 0.3 mol/L), which were suppressed by the TRPV4 blocker ruthenium red (RuR). Shear stress-induced vasodilatation was similarly blocked by RuR and also by the phospholipase A 2 inhibitor arachidonyl trifluoromethyl ketone (AACOCF 3 ). 4␣PDD produced endothelium-derived hyperpolarizing factor (EDHF)-type responses in A gracilis but not in rat carotid artery. Shear stress did not produce EDHF-type vasodilatation in either vessel type. Conclusions-Ca2ϩ entry through endothelial TRPV4 channels triggers NO-and EDHF-dependent vasodilatation. Moreover, TRPV4 appears to be mechanistically important in endothelial mechanosensing of shear stress. Key Words: endothelium-dependent vasodilatation Ⅲ transient receptor potential Ⅲ TRPV4 Ⅲ calcium Ⅲ shear stress Ⅲ nitric oxide Ⅲ 4␣PDD Ⅲ rat carotid artery C a 2ϩ -influx in response to mechanical or humoral stimulation plays a significant role in a variety of endothelial functions and especially in the Ca 2ϩ -dependent synthesis of endothelium-derived vasodilators such as NO, prostacyclin, or the endothelium-derived hyperpolarizing factor (EDHF). 1

Small-conductance (KCa2.1-2.3) and intermediate-conductance (KCa3.1) calcium-activated K ϩ channels are critically involved in modulating calcium-signaling cascades and membrane potential in both excitable and nonexcitable cells. Activators of these channels constitute useful pharmacological tools and potential new drugs for the treatment of ataxia, epilepsy, and hypertension. Here, we used the neuroprotectant riluzole as a template for the design of KCa2/3 channel activators that are potent enough for in vivo studies.

Immunohistochemical examination of lymph nodes may improve the pathological staging of esophageal cancer.

Background-It has been proposed that activation of endothelial SK3 (K Ca 2.3) and IK1 (K Ca 3.1) K ϩ channels plays a role in the arteriolar dilation attributed to an endothelium-derived hyperpolarizing factor (EDHF). However, our understanding of the precise function of SK3 and IK1 in the EDHF dilator response and in blood pressure control remains incomplete. To clarify the roles of SK3 and IK1 channels in the EDHF dilator response and their contribution to blood pressure control in vivo, we generated mice deficient for both channels. Methods and Results-Expression and function of endothelial SK3 and IK1 in IK1Ϫ/Ϫ /SK3 T/T mice was characterized by patch-clamp, membrane potential measurements, pressure myography, and intravital microscopy. Blood pressure was measured in conscious mice by telemetry. Combined IK1/SK3 deficiency in IK1Ϫ/Ϫ /SK3 T/T (ϩdoxycycline) mice abolished endothelial K Ca currents and impaired acetylcholine-induced smooth muscle hyperpolarization and EDHFmediated dilation in conduit arteries and in resistance arterioles in vivo. IK1 deficiency had a severe impact on acetylcholine-induced EDHF-mediated vasodilation, whereas SK3 deficiency impaired NO-mediated dilation to acetylcholine and to shear stress stimulation. As a consequence, SK3/IK1-deficient mice exhibited an elevated arterial blood pressure, which was most prominent during physical activity. Overexpression of SK3 in IK1Ϫ/Ϫ /SK3 T/T mice partially restored EDHF-and nitric oxide-mediated vasodilation and lowered elevated blood pressure. The IK1-opener SKA-31 enhanced EDHF-mediated vasodilation and lowered blood pressure in SK3-deficient IK1 ϩ/ϩ /SK3 T/T (ϩdoxy-cycline) mice to normotensive levels. Conclusions-Our study demonstrates that endothelial SK3 and IK1 channels have distinct stimulus-dependent functions, are major players in the EDHF pathway, and significantly contribute to arterial blood pressure regulation. Endothelial K Ca channels may represent novel therapeutic targets for the treatment of hypertension.