ISSN 1990-7478, Biochemistry (Moscow) Supplement Series A: Membrane and Cell Biology, 2009, Vol. 3, No. 1, pp. 71–80. © Pleiades Publishing, Ltd., 2009. Original Russian Text © R.A. Romanov, N.V. Kabanova, S.L. Malkin, S.S. Kolesnikov, 2009, published in Biologicheskie Membrany, 2009, Vol. 26, No. 1, pp. 64–74. 71 The cell population in taste bud is heterogeneous and includes four different types of cells identified according to morphological criteria. Three cell types (I, II, and III) are characterized by a spindle shape and involved in receptive, supporting, and secretory func- tions [1–3]. The rounded cells of type IV, or basal cells, are considered as precursor cells that differentiate into the taste cells of the first three types [3]. The mean life- time of type IV cells in the taste bud is about 10 days [4, 5]. The population of spindle cells is heterogenic not only morphologically; these cells can be subdivided into three groups (types A, B, C) according to their electrophysiological behavior [6]. Since electrophysio- logical properties of the taste cells strongly correlate with the expression profile of certain marker proteins [7, 8], we revealed the following association between the cells: type A = type II, type B = type III, and type C = type I [8]. Taste cells of type II and III possess voltage-gated (VG) Na + -channels [6–8], so these cells can generate action potential spikes [8–10]. As a rule, repolarization of electrically excitable cells occurs owing to activation of VG K + -channels of various types; therefore, in a great variety of cells the outwards currents are carried predominantly by potassium ions. At the same time, type II taste cell demonstrate pronounced VG outward currents up to several nanoamperes, which are pre- served during inhibition of usual potassium currents— for example, in experiments with dialysis of cells with a solution containing CsCl instead of KCl. Therefore, the aim of this study was to describe the ionic channels that account for the VG outwards currents with unusual features in type II taste cells. EXPERIMENTAL Experiments were carried out on single taste cells isolated from vallate and foliate papilla of mouse tongue of 6 to 12 week old mice. The isolation of taste cells was described elsewhere [11]. The electrical activity of taste cells was recorded by patch clamp technique in the whole-cell (WC) or perfo- rated patch mode. In these experiments we used an Axopatch 200B amplifier, a DigiData 1322A AD/DA converter, and a licensed software pClamp 8.0 (all from Axon Instruments, USA). The perfusion system allowed a rapid change of the extracellular solution at the rate of 0.1–1.0 ml/s [12]. The recording pipettes were filled with solutions containing (in mM): 140 KCl/CsCl, 1 MgCl 2 , 0.5 EGTA, 10 HEPES-KOH (CsOH), pH 7.4. In the case of perforated patch clamp, amphotericin B (400 μg/ml) was added to the pipette- filling solution. In whole-cell experiments (excluding some special experiments), the pipette solution con- tained 2 mM MgATP instead of 1 mM MgCl 2 . The standard extracellular solution contained (in mM): 140 NaCl, 2.5 KCl, 1 CaCl 2 , 1 MgCl 2 , 10 HEPES- NaOH, pH 7.4. When needed, the solutions were mod- ified by equimolar replacement of Na + and/or K + by other cations or of Cl – , by other anions. Fifty-mM solu- tion of Mg 2 ATP was prepared in the following way. Aqueous solution of MgATP at the concentration greater than 50 mM was titrated with Mg(OH) 2 to pH 7.4 and then the solution was filtered to remove the undissolved Mg(OH) 2 . Then the solution was diluted with water to reach the concentration of Mg 2 ATP equal to 50 mM. All salts, alkalies, EGTA, EDTA, Na 2 ATP , MgATP, and fluorescein were from Sigma-Aldrich (USA). Nonselective Voltage-Gated Ionic Channels in Type II Taste Cells R. A. Romanov, N. V. Kabanova, S. L. Malkin, and S. S. Kolesnikov Institute of Cell Biophysics, Russian Academy of Sciences, ul. Institutskaya, 3, Pushchino, Moscow oblast, 142290 Russia; e-mail: cheglok@rambler.ru Received October 6, 2008 Abstract—In cells of different types outward voltage-gated (VG) ion currents are generally carried by potas- sium ions. However, in mouse type II taste cells these currents persist when K + -selective ion channels are inhib- ited. In this study, we examined the ion channels that provide a pathway for atypical VG outward currents in type II taste cells. These channels are found to be weakly selective and permeabile to large molecules such as NMDG, gluconate, and ATP. According to non-stationary fluctuation analysis, single channel conductance is about 200 pS. The data obtained suggest that the nonselective ion channels are similar to hemichannels formed by connexins, the gap-junction proteins, in the plasma membrane of vertebrate cells. Key words: nonselective channels, voltage-gated channels, type II taste cells, connexins, ATP. DOI: 10.1134/S1990747809010103