Voa protein is a subunit of V-ATPase proton pump which is essential to acidify intracellular organelles including synaptic vesicles. Voa1 is one of the four isoforms of Voa family with strong expression in neurons. Our present study was aimed to examine the role of Voa1 protein in mammalian brain neurons. To circumvent embryonic lethality, we generated conditional Voa1 knockout mice in which Voa1 was selectively deleted from forebrain pyramidal neurons. We performed experiments in the Voa1 knockout mice of ages 5-6 months to assess the persistent effects of Voa1 deletion. We found that the Voa1 knockout mice exhibited poor performance in the Morris water maze test compared to control mice. In addition, synaptic field potentials of the hippocampal CA1 region were greatly diminished in the Voa1 knockout mice when examined in brain slices in vitro. Furthermore, brain histological experiments showed severe degeneration of dorsal hippocampal CA1 neurons while CA3 neurons were largely preserved. The CA1 neurodegeneration was associated with general brain atrophy as overall hemispheric areas were reduced in the Voa1 cKO mice. Despite the CA1 degeneration and dysfunction, electroencephalographic recordings from the hippocampal CA3 area revealed aberrant spikes and non-convulsive discharges in the Voa1 knockout mice but not in control mice. These hippocampal spikes were suppressed by single intra-peritoneal injection of diazepam which is a benzodiazepine GABAA receptor enhancer. Together these results suggest that Voa1 related activities are essential for the survival of the targeted neurons in the dorsal hippocampal CA1 as well as other forebrain areas. We postulate that the Voa1 knockout mice may serve as a valuable model for further investigation of V-ATPase dysfunction related neuronal degeneration and functional abnormalities in forebrain areas particularly the hippocampus.