In amyotrophic lateral sclerosis (ALS) hallmark symptoms include motor neurone hyperexcitability (Bostock et al. 1995; Zanette et al. 2002) raising the possibility that abnormal membrane conductance of motor neurones underlies the disease progression (Zanette et al. 2002). Furthermore, oxidative stress, a putative factor in ALS, causes irreversible depolarisation and enhanced calcium influx precipitating cell death (Herson & Ashford, 1997). Maintenance of the resting membrane potential (RMP) may prevent cell death and we describe here which channels in motor neurones may contribute to RMP.

Spinal cord cultures (islolated from E13 Balb-C mice, humanely killed) were grown as previously described (Boolaky et al. 2002). Motor neurones were identified by both morphology and immunohistochemistry. Whole-cell, excised inside-out and cell-attached recordings were made. The bath contained (mM): 140 NaCl; 3 KCl; 2 MgCl₂; 2 CaCl₂; 10 Hepes; 10 glucose; 1 µM TTX; buffered to pH 7.4 using NaOH. The pipette solution was (mM): 140 KCl; 5 NaCl; 1 MgCl₂; 1 CaCl₂; 11 EGTA; Hepes 10; buffered to pH 7.2 with KOH. In some experiments 10 mM TEA was added to the pipette.

In whole-cell measurements motor neurones have healthy RMPs (-57 ± 11 mV; mean ± S.D., n = 19). Single channels were present in 45 % of the cells. The predominant channel at 0 mV pipette potential (PP) was of small conductance, the mean inward conductance being 46 ± 11 pS, the current reversing at a PP of -57 ± 21 mV (n = 25). At RMP the mean unitary current was -2.6 ± 0.6 pA (n = 13). The open probability (P_o) was voltage independent. An intermediate conductance channel (149 ± 9 pS) was present in 23 % of cells. The channel current reversed at a PP of -67 ± 25 mV (n = 10) and the activity was voltage independent. A large TEA-sensitive channel with a mean conductance of 203 ± 28 pS (n = 6) was present in cell-attached patches and in excised inside-out patches (169 ± 34 pS, n = 6). The channel current showed inward rectification in asymmetrical K⁺ (reverse gradient). This channel was both voltage and calcium dependent, the P_o increasing with depolarisation and increased ‘intracellular’ calcium.

Three channels which may contribute to resting conductance have been identified in motor neurones in culture. The large conductance channel is clearly the maxi-K potassium channel. The effects of glutamate excitotoxicity and oxidative stress on these channels is being investigated.