Hereditary canine spinal muscular atrophy (HCSMA) is an autosomally dominant disease of motor neurons that shares many pathological features with human motor neuron disease. A particularly striking feature of the affected, accelerated phenotype (homozygous HCSMA) is that profound weakness develops before appreciable motor neuron cell death occurs (Cork et al., 1989a), implying that motor unit functional defects occur initially. The purpose of this study was to identify the site of these defects and characterize their nature. In most young homozygotes (2–3 months postnatal), motor neurons were encountered that could support orthodromic action potential propagation to the muscle but did not activate muscle fibers. The tetanic forces of innervated motor units in young homozygotes tended to be smaller than those in closely age-matched clinically normal animals. In older homozygotes (approximately 4.5 months, postnatal), all motor neurons sampled were capable of activating muscle fibers, but many motor units displayed abnormal behavior including an inability to sustain force output during high frequency activation. Motor units exhibiting tetanic failure also showed proportionately greater twitch potentiation than nonfailing units of similar unpotentiated twitch amplitude. Tetanic failure and large potentiation tended to occur in motor units that possessed the slowest contraction speeds. These results indicate that motor neuron functional defects in HCSMA appear initially in the most distal parts of the motor axon and involve defective neurotransmission. The possible roles of distal nerve degeneration, motor terminal sprouting, and synaptic transmission in causing these deficits are considered.