measurement-ion-concentration-unstirred-boundary-layer-with-open

Measurement of Ion Concentration in the Unstirred Boundary Layer with Open Patch-Clamp Pipette: Implications in Control of Ion Channels by Fluid Flow

Fluid flow is an important environmental stimulus that controls many physiological and pathological processes, such as fluid flow-induced vasodilation. ...

Konkuk University School of Medicine

Activation of L-type voltage-dependent Ca channels (VDCC) by membrane stretch contributes to many biological responses such as myogenic contraction of arteries. However, mechanism for the stretch-induced VDCC activation is unclear. In this study, we examined the hypothesis that caveolar remodeling and its related signaling cascade contribute to the stretch-induced activation of VDCC in rat mesenteric arterial smooth muscle cells. The VDCC currents were recorded with nystatin-perforated or with conventional whole-cell patch-clamp technique. Hypotonic (~230 mOsm) swelling-induced membrane stretch reversibly increased the VDCC currents. Electron microscope and confocal imaging analysis revealed that both hypotonic swelling and cholesterol depletion by methyl-β-cychlodextrin (MβCD) similarly disrupted the caveolae structure and translocated caveolin-1 (Cav-1) from membrane to cytosolic space. Accordingly, MβCD also increased VDCC currents. Moreover, subsequent hypotonic swelling after MβCD treatment failed to increase the VDCC currents further. Western blotting experiments revealed that hypotonic swelling phosphorylated Cav-1 and JNK. Inhibitors of tyrosine kinases (genistein) and JNK (SP00125) prevented the swelling-induced facilitation of VDCC currents. Knockdown of Cav-1 by small interfering RNA blocked both the VDCC current facilitation by stretch and the related phosphorylation of JNK. Taken together, the results suggest that membrane stretch is transduced to the facilitation of VDCC currents via caveolar structure-dependent tyrosine phosphorylation of Cav-1 and subsequent activation of JNK in rat mesenteric arterial myocytes.

Caveolar remodeling is a critical mechanotransduction mechanism of the stretch-induced L-type Ca channel activation in vascular myocytes.

Mulberry (Morus alba) has been used in traditional oriental medicine since ages. Recently, it has been reported that mulberry produces hypotensive effects through the eNOS signaling pathway. However, the mechanism underlying the hypotensive effects of mulberry is not entirely clear. Moreover, the effects of mulberry on vascular remodeling events such as hyperplasia, an important etiology in the pathogenesis of hypertension and arteriosclerosis, are also ambiguous. Here, we hypothesized that an ethanolic extract of mulberry fruit (EMF) has beneficial effects on vascular remodeling and produces hypotensive effects. The effects of a 6-week oral administration of EMF were examined in spontaneously hypertensive rats (SHRs). The animals were divided into four groups: normotensive control (Wistar Kyoto rats), non-treated SHR, low-dose (100 mg/kg) EMF-treated SHR, and high-dose (300 mg/kg) EMF-treated SHR. Our results showed that the EMF-diet normalizes hypertension in SHRs in a dose-dependent manner, by preventing smooth muscle proliferation, thickening of the tunica media, and vascular hyper-reactivity. The endothelial functions were not substantially affected by the EMF diet in our experimental setting. In conclusion, we suggest that the mulberry fruit could act as a food supplement for reducing blood pressure in hypertensive subjects through its effects on smooth muscle proliferation and vascular contractility.

Effects of an ethanolic extract of mulberry fruit on blood pressure and vascular remodeling in spontaneous hypertensive rats.

Oxidative stress is associated with many cardiovascular diseases, such as hypertension and arteriosclerosis. Oxidative stress reportedly activates the L-type voltage-gated calcium channel (VDCC) and elevates [Ca] in many cells. However, how oxidative stress activates VDCC under clinical setting and the consequence for arteries are unclear. Here, we examined the hypothesis that hydrogen peroxide (HO) regulates membrane potential (Em) by altering Na influx through cation channels, which consequently activates VDCC to induce vasoconstriction in rat mesenteric arteries. To measure the tone of the endothelium-denuded arteries, a conventional isometric organ chamber was used. Membrane currents and Em were recorded by the patch-clamp technique. [Ca] and [Na] were measured with microfluorometry using Fura2-AM and SBFI-AM, respectively. We found that HO (10 and 100 µM) increased arterial contraction, and nifedipine blocked the effects of HO on isometric contraction. HO increased [Ca] as well as [Na], and depolarized Em. Gd (1 µM) blocked all these HO-induced effects including Em depolarization and increases in [Ca] and [Na]. Although both nifedipine (30 nM) and low Na bath solution completely prevented the HO-induced increase in [Na], they only partly inhibited the HO-induced effects on [Ca] and Em. Taken together, the results suggested that HO constricts rat arteries by causing Em depolarization and VDCC activation through activating Gd-and nifedipine-sensitive, Na-permeable channels as well as Gd-sensitive Ca-permeable cation channels. We suggest that unidentified Na-permeable cation channels as well as Ca-permeable cation channels may function as important mediators for oxidative stress-induced vascular dysfunction.

Kyung Chul Shin

Department of Physiology,
KU Open Innovation Center,
Research Institute of Medical Science,
Department of Physiology, KU Open Innovation Center, Research Institute of Medical Science

Konkuk University School of Medicine

Measurement of Ion Concentration in the Unstirred Boundary Layer with Open Patch-Clamp Pipette: Implications in Control of Ion Channels by Fluid Flow

Kyung Chul Shin

Department of Physiology,KU Open Innovation Center,Research Institute of Medical Science,Department of Physiology, KU Open Innovation Center, Research Institute of Medical Science

Konkuk University School of Medicine

Measurement of Ion Concentration in the Unstirred Boundary Layer with Open Patch-Clamp Pipette: Implications in Control of Ion Channels by Fluid Flow

Department of Physiology,
KU Open Innovation Center,
Research Institute of Medical Science,
Department of Physiology, KU Open Innovation Center, Research Institute of Medical Science