In the current study, we analyzed the impact of antenatal betamethasone on macroscopic cerebellar development, Purkinje cell morphology and the expression of the neuroprotective protein calbindin-D28k. Pregnant rats (Sprague-Dawley) were randomly divided into two experimental groups: control (CONT) and betamethasone-treated (BET). At gestational day 20 (G20), BET dams were subcutaneously injected with a solution of 0.17 mg kg⁻¹ of betamethasone, while CONT animals received a similar volume of saline. At postnatal days 22 (P22) and P52, BET and CONT offspring were behaviorally evaluated, and the cerebella were histologically and immunohistochemically processed. Animals that were prenatally treated with a single course of betamethasone exhibited long-lasting behavioral changes consistent with anxiety-like behavior in the open-field test, together with (1) reduced cerebellar weight and volume, (2) Purkinje cell dendritic atrophy, and (3) an overexpression of calbindin-D28k. The current results indicate that an experimental single course of betamethasone in pregnant rats produces long-lasting anxiety-like behaviors, together with macroscopic and microscopic cerebellar alterations.