Survival of Swiss-Webster mouse cerebellar granule neurons is promoted by a combination of potassium channel blockers

<p>Cultured cerebellar granule neurons (CGN) are commonly used to assess neurotoxicity, but are routinely maintained in supraphysiological (25 mM) extracellular K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> concentrations [K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup>]<sub style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">o</sub>. We investigated the effect of potassium channel blockade on survival of CGN derived from Swiss-Webster mice in supraphysiological (25 mM) and physiological (5.6 mM) [K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup>]<sub style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">o</sub>. CGN were cultured for 5 days in 25 mM K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup>, then in 5.6 mM K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> or 25 mM K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> (control). Viability, assayed 24 h later by 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) reduction and by lactate dehydrogenase (LDH) release, was ∼50% in 5.6 mM K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> versus 25 mM K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> (<em>p</em> < .001). Potassium channel blockers, 2 mM 4-aminopyridine (4-AP), 2 mM tetraethylammonium (TEA) or 1 mM Ba<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">2+</sup>, individually afforded limited protection in 5.6 mM K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup>. However, survival in 5.6 mM K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> with a combination of 4-AP, TEA and Ba<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">2+</sup> was similar to survival in 25 mM K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> without blockers (<em>p</em> < .001 versus 5.6 mM K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> alone). CGN survival in 25 mM K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> was attenuated 25% by 2 μM nifedipine (<em>p</em> > .001), but nifedipine did not attenuate neuroprotection by K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> channel blockers. Together, these results suggest that the survival of CGN depends on the K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> permeability of the membrane rather than the activity of a particular type of K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> channel, and that the mechanism of neuroprotection by K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup> channel blockers is different from that of elevated [K<sup style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">+</sup>]<sub style="border: 0px; font-size: 0.75em; margin: 0px; padding: 0px; line-height: 0; color: rgb(46, 46, 46); font-family: 'Arial Unicode MS', 'Arial Unicode', Arial, 'URW Gothic L', Helvetica, Tahoma, sans-serif; text-align: justify; word-spacing: -1.03587377071381px; background-color: rgb(255, 255, 255);">o</sub>.</p>