A membrane potential-sensitive dye for vascular smooth muscle cells assays.
Changes in membrane potential of rat aorta smooth muscle cells were investigated using the bis-oxonol sensitive probe DIBAC2(3). We compared the changes in membrane potential induced by a high external KCl concentration in aorta smooth muscle cells from normotensive 2 kidney (2K) and from renal hypertensive 2 kidney-1 clip (2K-1C) rats. The spectral properties of the membrane potential were first characterized in aqueous buffers and in cultured smooth muscle cells from 2K and 2K-1C rat aortas. Fluorescence emission and the images were recorded using a laser scanning confocal microscope. The relationship between fluorescence intensity (FI) and membrane potential (psi(m)) as a function of the increasing extracellular KCl concentration was linear in the 5-40 mmol/L KCl range in both 2K and 2K-1C rat aorta cells. Cell membranes from 2K-1C rat aorta cells were more depolarized (-55 mV) than 2K rat aorta cells (-65 mV). The results show that in 2K-1C aorta cells only 10 mmol/L KCl was needed to induce complete membrane depolarization while in 2K cells 40 mmol/L KCl was needed to induce a similar effect. This study clearly shows that the method is suitable to measure the membrane potential in cultured smooth muscle cells.
Sguilla
Fabiana S
FS
Laboratório de Farmacologia, Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Av. do Café, s/no. 14040-903 Ribeirão Preto, SP, Brazil.
Tedesco
Antonio C
AC
Bendhack
Lusiane M
LM
D000818:Animals; D001011:Aorta / Q000033:anatomy & histology; D001794:Blood Pressure / Q000502:physiology; D002138:Calibration; D005456:Fluorescent Dyes / Q000737:chemistry / Q000378:metabolism*; D006973:Hypertension / Q000378:metabolism; D008297:Male; D008564:Membrane Potentials / Q000502:physiology*; D018613:Microscopy, Confocal; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D017208:Rats, Wistar; D013858:Thiobarbiturates / Q000737:chemistry / Q000378:metabolism*
D005456:Fluorescent Dyes; D013858:Thiobarbiturates; C025284:bis(1,3-diethylthiobarbiturate)trimethineoxonol