Large conductance voltage- and calcium-activated potassium (BKCa) channels link electrical excitability with cellular signalling. Multiple conserved sites on the c-terminus of the α-subunit confer regulation by divalent cations such as magnesium, which can alter both single-channel conductance and open probability (Po) (Yang et al. 2006). However, alternative splicing of the α-subunit can alter BKCa channel properties including sensitivity to divalent cations. Here we investigated the ability of magnesium ions to modulate the single-channel properties of an alternatively spliced BKCa channel variant, the stress regulated exon (STREX). In addition, as the monovalent lithium ion shares many physicochemical properties, and interacts in a competitive manner, with magnesium (Mota de Freitas et al. 2006) the effects of lithium were also investigated. Single-channel patch clamp recording was used to determine the activity of the STREX BKCa channel variant expressed in HEK 293 cells. Excised inside-out patch recordings were made in symmetrical 140mM KCl, using pipettes with resistances of 5-15MΩ. The intracellular free calcium ion concentration was buffered using BAPTA to 0.1 or 1.0 µM. In excised inside out patches, when the concentration of free magnesium or free lithium at the intracellular surface of the membrane was raised from 2mM to 6mM the STREX variant single-channel amplitude was reduced by 30% and 25% respectively. For example, at 40mV the channel amplitude fell from 10.5 ± 0.1 pA (n=13) to 7.5 ± 0.1 pA in magnesium (n=4) and 7.7 ± 0.4 pA in lithium (n=5) (data are mean ± s.e.m., p<0.0001 ANOVA plus Tukey post-hoc). This was accompanied by a significant increase in Po from 0.21 ± 0.01 (n=13) to 0.43 ± 0.03 for magnesium (n=4) at 40mV (p<0.0001 Student's paired t-test). In contrast, lithium had no significant effect on STREX variant Po. The effects of both ions were voltage dependent in that the decrease in channel amplitude was only observed at depolarized potentials (+20mV to +80mV), and were reversible upon washout. Control experiments for lithium in which the monovalent ion sodium was increased from 2mM to 6mM did not alter either the Po or single-channel conductance of the STREX variant. The data indicate that the STREX BKCa channel variant is sensitive to modulation by both magnesium and lithium ions. However, the ions display a difference in their ability to modulate conductance and Po, which may be indicative of separate sites of action. Further work, including site directed mutagenesis will hopefully resolve these sites.