Tags

Type your tag names separated by a space and hit enter

Properties of HERG channels stably expressed in HEK 293 cells studied at physiological temperature.
Biophys J. 1998 Jan; 74(1):230-41.BJ

Abstract

We have established stably transfected HEK 293 cell lines expressing high levels of functional human ether-a go-go-related gene (HERG) channels. We used these cells to study biochemical characteristics of HERG protein, and to study electrophysiological and pharmacological properties of HERG channel current at 35 degrees C. HERG-transfected cells expressed an mRNA band at 4.0 kb. Western blot analysis showed two protein bands (155 and 135 kDa) slightly larger than the predicted molecular mass (127 kDa). Treatment with N-glycosidase F converted both bands to smaller molecular mass, suggesting that both are glycosylated, but at different levels. HERG current activated at voltages positive to -50 mV, maximum current was reached with depolarizing steps to -10 mV, and the current amplitude declined at more positive voltages, similar to HERG channel current expressed in other heterologous systems. Current density at 35 degrees C, compared with 23 degrees C, was increased by more than twofold to a maximum of 53.4 +/- 6.5 pA/pF. Activation, inactivation, recovery from inactivation, and deactivation kinetics were rapid at 35 degrees C, and more closely resemble values reported for the rapidly activating delayed rectifier K+ current (I(Kr)) at physiological temperatures. HERG channels were highly selective for K+. When we used an action potential clamp technique, HERG current activation began shortly after the upstroke of the action potential waveform. HERG current increased during repolarization to reach a maximum amplitude during phases 2 and 3 of the cardiac action potential. HERG contributed current throughout the return of the membrane to the resting potential, and deactivation of HERG current could participate in phase 4 depolarization. HERG current was blocked by low concentrations of E-4031 (IC50 7.7 nM), a value close to that reported for I(Kr) in native cardiac myocytes. Our data support the postulate that HERG encodes a major constituent of I(Kr) and suggest that at physiological temperatures HERG contributes current throughout most of the action potential and into the postrepolarization period.

Authors+Show Affiliations

Department of Medicine (Cardiology), University of Wisconsin, Madison 53792, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Comparative Study
Journal Article

Language

eng

PubMed ID

9449325

Citation

Zhou, Z, et al. "Properties of HERG Channels Stably Expressed in HEK 293 Cells Studied at Physiological Temperature." Biophysical Journal, vol. 74, no. 1, 1998, pp. 230-41.
Zhou Z, Gong Q, Ye B, et al. Properties of HERG channels stably expressed in HEK 293 cells studied at physiological temperature. Biophys J. 1998;74(1):230-41.
Zhou, Z., Gong, Q., Ye, B., Fan, Z., Makielski, J. C., Robertson, G. A., & January, C. T. (1998). Properties of HERG channels stably expressed in HEK 293 cells studied at physiological temperature. Biophysical Journal, 74(1), 230-41.
Zhou Z, et al. Properties of HERG Channels Stably Expressed in HEK 293 Cells Studied at Physiological Temperature. Biophys J. 1998;74(1):230-41. PubMed PMID: 9449325.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Properties of HERG channels stably expressed in HEK 293 cells studied at physiological temperature. AU - Zhou,Z, AU - Gong,Q, AU - Ye,B, AU - Fan,Z, AU - Makielski,J C, AU - Robertson,G A, AU - January,C T, PY - 1998/2/4/pubmed PY - 1998/2/4/medline PY - 1998/2/4/entrez SP - 230 EP - 41 JF - Biophysical journal JO - Biophys J VL - 74 IS - 1 N2 - We have established stably transfected HEK 293 cell lines expressing high levels of functional human ether-a go-go-related gene (HERG) channels. We used these cells to study biochemical characteristics of HERG protein, and to study electrophysiological and pharmacological properties of HERG channel current at 35 degrees C. HERG-transfected cells expressed an mRNA band at 4.0 kb. Western blot analysis showed two protein bands (155 and 135 kDa) slightly larger than the predicted molecular mass (127 kDa). Treatment with N-glycosidase F converted both bands to smaller molecular mass, suggesting that both are glycosylated, but at different levels. HERG current activated at voltages positive to -50 mV, maximum current was reached with depolarizing steps to -10 mV, and the current amplitude declined at more positive voltages, similar to HERG channel current expressed in other heterologous systems. Current density at 35 degrees C, compared with 23 degrees C, was increased by more than twofold to a maximum of 53.4 +/- 6.5 pA/pF. Activation, inactivation, recovery from inactivation, and deactivation kinetics were rapid at 35 degrees C, and more closely resemble values reported for the rapidly activating delayed rectifier K+ current (I(Kr)) at physiological temperatures. HERG channels were highly selective for K+. When we used an action potential clamp technique, HERG current activation began shortly after the upstroke of the action potential waveform. HERG current increased during repolarization to reach a maximum amplitude during phases 2 and 3 of the cardiac action potential. HERG contributed current throughout the return of the membrane to the resting potential, and deactivation of HERG current could participate in phase 4 depolarization. HERG current was blocked by low concentrations of E-4031 (IC50 7.7 nM), a value close to that reported for I(Kr) in native cardiac myocytes. Our data support the postulate that HERG encodes a major constituent of I(Kr) and suggest that at physiological temperatures HERG contributes current throughout most of the action potential and into the postrepolarization period. SN - 0006-3495 UR - https://www.unboundmedicine.com/medline/citation/9449325/Properties_of_HERG_channels_stably_expressed_in_HEK_293_cells_studied_at_physiological_temperature_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0006-3495(98)77782-3 DB - PRIME DP - Unbound Medicine ER -