Parvalbumin-expressing neurons are a type of inhibitory intermediate neuron that play an important role in terminating seizures. The aim of the present study was to use lentiviral construction and packaging technology to overexpress and silence the parvalbumin gene in pheochromocytoma (PC12) cells, and to evaluate how parvalbumin influences the metabolic pathway involving glutamate and γ-aminobutyric acid (GABA). In this work, Immunofluorescence staining was used to verify the differentiation of PC12 cells into neurons after adding nerve growth factor (NGF). Western blotting and real-time quantitative polymerase chain reaction (qRT-PCR) were used to confirm lentivirus-mediated knockdown or overexpression of parvalbumin. Expression of parvalbumin, the 65-kDa GAD isoform (GAD65), and the 67-kDa GAD isoform (GAD67) in neuronal cells was examined at the mRNA and protein levels using qRT-PCR, western blotting and immunofluorescence staining, while intracellular glutamate and GABA levels were determined by high performance liquid chromatography (HPLC). We demonstrate that the expression of parvalbumin is associated with GAD65 and GAD67. Interestingly, overexpression of parvalbumin up-regulated GAD65 and GAD67, increased GABA concentration, and decreased glutamate concentration. Silencing of parvalbumin led to the opposite effects. Altogether, parvalbumin affected the expression of GAD65 and GAD67, thereby influencing the metabolic pathway involving glutamate and GABA.