Guinea pig horizontal cells express GABA, the GABA-synthesizing enzyme GAD₆₅, and the GABA vesicular transporter

γ-Aminobutyric acid (GABA) is likely expressed in horizontal cells of all species, although conflicting physiological findings have led to considerable controversy regarding its role as a transmitter in the outer retina. This study has evaluated key components of the GABA system in the outer retina of guinea pig, an emerging retinal model system. The presence of GABA, its ratelimiting synthetic enzyme glutamic acid decarboxylase (GAD₆₅ and GAD₆₇ isoforms), the plasma membrane GABA transporters (GAT-1 and GAT-3), and the vesicular GABA transporter (VGAT) was evaluated by using immunohistochemistry with well-characterized antibodies. The presence of GAD₆₅ mRNA was also evaluated by using laser capture microdissection and reverse transcriptase-polymerase chain reaction. Specific GABA, GAD ₆₅, and VGAT immunostaining was localized to horizontal cell bodies, as well as to their processes and tips in the outer plexiform layer. Furthermore, immunostaining of retinal whole mounts and acutely dissociated retinas showed GAD₆₅ and VGAT immunoreactivity in both A-type and B-type horizontal cells. However, these cells did not contain GAD₆₇, GAT-1, or GAT-3 immunoreactivity. GAD₆₅ mRNA was detected in horizontal cells, and sequencing of the amplified GAD₆₅ fragment showed approximately 85% identity with other mammalian GAD₆₅ mRNAs. These studies demonstrate the presence of GABA, GAD₆₅, and VGAT in horizontal cells of the guinea pig retina, and support the idea that GABA is synthesized from GAD₆₅, taken up into synaptic vesicles by VGAT, and likely released by a vesicular mechanism from horizontal cells.