ROMK1 (Kir1.1) causes apoptosis and chronic silencing of hippocampal neurons

Lentiviral vectors were constructed to express the weakly rectifying kidney K+ channel ROMK1 (Kir1.1), either fused to enhanced green fluorescent prot ein (EGFP) or as a bicistronic message (ROMK1-CITE-EGFP). The channel was s tably expressed in cultured rat hippocampal neurons. Infected cells were ma intained for 2-4 wk without decrease in expression level or evidence of vir al toxicity, although 15.4 mM external KCl was required to prevent apoptosi s of neurons expressing functional ROMK1. No other trophic agents tested co uld prevent cell death, which was probably caused by K+ loss. This cell dea th did not occur in glia, which were able to support ROMK1 expression indef initely. Functional ROMK1, quantified as the nonnative inward current at -1 44 mV in 5.4 mM external K+ blockable by 500 mu M Ba2+, ranged from 1 to 40 pA/pF. Infected neurons exhibited a Ba2+-induced depolarization of 7 +/- 2 mV relative to matched EGFP-infected controls, as well as a 30% decrease i n input resistance and a shift in action potential threshold of 2.6 +/- 0.5 mV. This led to a shift in the relation between injected current and firin g frequency, without changes in spike shape, size, or timing. This shift, w hich quantifies silencing as a function of ROMK1 expression, was predicted from Hodgkin-Huxley models. No cellular compensatory mechanisms in response to expression of ROMK1 were identified, making ROMK1 potentially useful fo r transgenic studies of silencing and neurodegeneration, although its letha lity in normal K+ has implications for the use of K+ channels in gene thera py.