Suppression of the rat microglia Kv1.3 current by src-family tyrosine kinases and oxygen/glucose deprivation

Microglia activate following numerous acute insults to the brain, including oxygen/glucose deprivation (OGD), and both protein tyrosine kinases (PTKs) and K+ channels have been implicated in their activation. We identified Kv 1.3 (voltage-gated potassium channel) protein in cultured rat microglia and confirmed that the native current is biophysically and pharmacologically s imilar to Kv1.3. To explore whether src-family PTKs regulate the microglial Ky current, we first heterologously expressed Kv1.3 in a microglia-like ce ll line derived from neonatal rat brain (MLS-9). The resulting large Kv1.3 current was eliminated by co-transfecting the constitutively active PTK, v- src, then rapidly restored by the PTK inhibitor, lavendustin A. Acute activ ation of endogenous src kinases by a peptide activator significantly reduce d the current, an effect that was mimicked by OGD. Similarly, in primary cu ltures of rat microglia, the endogenous Kv1.3-like current was inhibited by activating endogenous src-family PTKs and by OGD. Biochemical analysis sho wed that OGD increased the tyrosine phosphorylation of native Kv1.3 protein , which was alleviated by PTK inhibitors or reactive oxygen species (ROS) s cavengers. Conversely, the basal level of Kv1.3 phosphorylation was decreas ed by PTK inhibitors or scavengers of ROS. Together, our results point to a post-insertional downregulation of the microglial Kv1.3-like current by ox idative stress and tyrosine phosphorylation. This interaction may be facili tated by a multiprotein complex because, in cultured microglia, the endogen ous Kv1.3 and src proteins both bind to the scaffolding protein, post-synap tic density protein 95 (PSD-95). By associating with, and phosphorylating K v1.3, src is well positioned to regulate microglial responses to oxidative stress.