Exposure of lentiviral vectors to subneutral pH shifts the tropism from Purkinje cell to Bergmann glia

Abstract

Cerebellar Purkinje cells play an important role in cerebellar function; lesions of Purkinje cells result in the disruption of motor coordination and motor learning. Although selective gene delivery to Purkinje cells would be a powerful technique for the study of pathophysiology in the cerebellum, a method for such a delivery has not yet been established. Here we employed human immunodeficiency virus-derived lentiviral vectors pseudotyped with vesicular stomatitis virus glycoprotein to transduce Purkinje cells and examined factors that critically affect the viral tropism for Purkinje cells. Viral vectors encoding GFP were generated using different protocols, and were then injected into the mouse cerebellum. At 7 days and 2 months post-transduction, the relative proportions of transduced Purkinje cells were determined. Lentiviral vectors harvested from a medium of pH 7.2 preferentially transduced Purkinje cells (about half of the transduced cells). In contrast, when the viral vector was harvested from medium of <or= pH 7.0, only 12-26% of transduced cells were identified as Purkinje cells and 68-77% as Bergmann glia. A similar decrease in the efficiency of transduction for Purkinje cells, depending on the pH of the medium at the viral harvest, was observed in dissociated cell cultures. These results indicate that lentivector tropism for Purkinje cells is extremely sensitive to pH: a subtle decrease in the pH of the medium at the harvest shifts viral tropism strikingly towards Bergmann glia.