The number of ionotropic receptors in synapses is an essential factor for determining the efficacy of fast transmission. We estimated the number of functional AMPA receptors at single postsynaptic sites by a combination of two-photon uncaging of glutamate and the nonstationary fluctuation analysis in immature rat Purkinje cells (PCs), which receive a single type of excitatory input from climbing fibers. Areas of postsynaptic membrane specialization at the recorded synapses were measured by reconstruction of serial ultrathin sections. The number of functional AMPA receptors was proportional to the synaptic area with a density of approximately 1280 receptors/microm2. Moreover, highly sensitive freeze-fracture replica labeling revealed a homogeneous density of immunogold particles for AMPA receptors in synaptic sites (910 +/- 36 particles/microm2) and much lower density in extrasynaptic sites (19 +/- 2 particles/microm2) in the immature PCs. Our results indicate that in this developing synapse, the efficacy of transmission is determined by the synaptic area.