Temperature-dependent expression of a squid Kv1 channel in Sf9 cells and functional comparison with the native delayed rectifier

SqKv1A is a cDNA that encodes a Kv1 (Shaker-type) alpha -subunit expressed only in the giant axon and the parental giant fiber lobe (GFL) neurons of t he squid stellate ganglion. We incorporated SqKv1A into a recombinant bacul ovirus for expression in the insect Sf9 cell line. Whale-cell patch-clamp r ecordings reveal that very few cells display functional potassium current ( I,) if cultured at the standard postinfection temperature of 27 degreesC, A t 18 degreesC, less SqKv1A protein is produced than at 27 degreesC, but cel ls with I-K currents are much more numerous and can survive for at least 20 days postinfection (vs. similar to5 days at 27 degreesC). Activation and d eactivation kinetics of SqKv1A in Sf9 cells are slower (similar to3- and 10 -fold, respectively! than those of native channels in GFL neurons, but have similar voltage dependencies, The two cell types show only subtle differen ces in steady-state voltage-dependence of conductance and inactivation. Rat es of I-K inactivation in 20 mM external K are identical in the two cell ty pes, but the sensitivity of inactivation to external tetraethylammonium (TE A) and K ions differ: inactivation of SqKv1A in Sf9 cells is slowed by exte rnal TEA and K ions, whereas inactivation of GFL I-K is largely insensitive . Functional differences are discussed in terms of factors that may be spec ific to cell-type, including the presence of presently unidentified Kv1 sub units in GFL neurons that might form heteromultimers with SqKv1A.