Here we describe two families with retinitis pigmentosa, a hereditary neurodegeneration of rod and cone photoreceptors in the retina. Affected family members were homozygous for loss-of-function mutations in IDH3B, encoding the beta-subunit of NAD-specific isocitrate dehydrogenase (NAD-IDH, or IDH3), which is believed to catalyze the oxidation of isocitrate to alpha-ketoglutarate in the citric acid cycle. Cells from affected individuals had a substantial reduction of NAD-IDH activity, with about a 300-fold increase in the K(m) for NAD. NADP-specific isocitrate dehydrogenase (NADP-IDH, or IDH2), an enzyme that catalyzes the same reaction, was normal in affected individuals, and they had no health problems associated with the enzyme deficiency except for retinitis pigmentosa. These findings support the hypothesis that mitochondrial NADP-IDH, rather than NAD-IDH, serves as the main catalyst for this reaction in the citric acid cycle outside the retina, and that the retina has a particular requirement for NAD-IDH.