The septal region of the brain consists of a heterogeneous population of GABAergic neurons that play an important role in the generation of hippocampal theta rhythms. While GABAergic neurons employ two isoforms of the enzyme glutamic acid decarboxylase (GAD) for the synthesis of GABA, distribution of GAD isoforms has not been investigated in the septum. Immunohistochemical techniques were used to investigate the expression of GAD enzymes in medial and lateral septum. GAD65 and GAD67 immunohistochemistry revealed dense fibers and punctuated immunoreactivity in septal regions. While few GAD65-positive neuronal somas were detected in medial septum, a significantly higher number of immunoreactive neurons were detected in lateral septum. GAD65- and GAD67-positive neurons in the lateral septum exhibit higher complexity of dendritic arborizations than in the medial septum where staining was mainly restricted to the soma. Presumptive axon terminals (puncta) showed abundant immunoreactivity predominantly for GAD65 isoforms in all septal regions. This suggests that septal GABAergic neurons differentially express GAD enzymes thereby potentially reflecting functional differences. Differences found between medial and lateral septal GABAergic neuronal populations are in agreement with the concept that medial and lateral septum are brain structures with highly different connectivity and function despite anatomical proximity.