The autonomic innvervation of the interstitial gland of the rat ovary was studied on days 4, 6, 10, 14 and 18 of pregnancy with the acetylcholinesterase procedure, the Falck-Hillarp technique and electron microscopy after 5-hydroxydopamine treatment. Acetylcholinesterase-positive nerves were present as perivascular plexuses at all stages studied. Adrenergic nerves were present in the interstitial gland in all stages studied. The number and intensity of interstitial fluorescent adrenergic nerves increased as pregnancy progressed. Measurement of norepinephrine with the fluorometric procedure showed a highly significant (p < 0.05) increase in the neurotransmitter in the ovary on days 14 and 18 as compared to day 4. Fine-structural studies after administration of the false transmitter, 5-hydroxydopamine, showed that the innervation of the steroidogenic cells of the interstitial gland is adrenergic.