Quantification of the functional expression of the Ca²⁺-activated K⁺channel K_Ca3.1 on microglia from adult human neocortical tissue

The K_Ca 3.1 channel (KCNN4) is an important modulator of microglia responses in rodents, but no information exists on functional expression on microglia from human adults. We isolated and cultured microglia (max 1% astrocytes, no neurons or oligodendrocytes) from neocortex surgically removed from epilepsy patients and employed electrophysiological whole-cell measurements and selective pharmacological tools to elucidate functional expression of K_Ca 3.1. The channel expression was demonstrated as a significant increase in the voltage-independent current by NS309, a K_Ca 3.1/K_Ca 2 activator, followed by full inhibition upon co-application with NS6180, a highly selective K_Ca 3.1 inhibitor. A major fraction (79%) of unstimulated human microglia expressed K_Ca 3.1, and the difference in current between full activation and inhibition (ΔK_Ca 3.1) was estimated at 292 ± 48 pA at -40 mV (n = 75), which equals at least 585 channels per cell. Serial K_Ca 3.1 activation/inhibition significantly hyperpolarized/depolarized the membrane potential. The isolated human microglia were potently activated by lipopolysaccharide (LPS) shown as a prominent increase in TNF-α production. However, incubation with LPS neither changed the K_Ca 3.1 current nor the fraction of K_Ca 3.1 expressing cells. In contrast, the anti-inflammatory cytokine IL-4 slightly increased the K_Ca 3.1 current per cell, but as the membrane area also increased, there was no significant change in channel density. A large fraction of the microglia also expressed a voltage-dependent current sensitive to the K_Ca 1.1 modulators NS1619 and Paxilline and an inward-rectifying current with the characteristics of a K_ir channel. The high functional expression of K_Ca 3.1 in microglia from epilepsy patients accentuates the need for further investigations of its role in neuropathological processes. GLIA 2016;64:2065-2078.