Diverse protocadherins (Pcdhs), which are encoded as a large cluster (composed of α, β and γ clusters) in the genome, are localized to axons and synapses. The Pcdhs have been proposed to contribute to the generation of sophisticated neural networks and to regulate brain function. To address the molecular roles of Pcdhs in regulating individual behavior, here we generated knockdown mice of Pcdh-α proteins and examined their behavioral abnormalities. There are two alternative splicing variants of the Pcdh-α constant region, Pcdh-α A and B isoforms, with different cytoplasmic tails. Pcdh-αΔBneo/ΔBneo mice, in which the Pcdh-α B splicing variant was absent and the Pcdh-α A isoforms were down-regulated to approximately 20% of the wild-type level, exhibited enhanced contextual fear conditioning and disparities in an eight-arm radial maze. Similar abnormalities were found in Pcdh-αΔAneo/ΔAneo mice, which lacked 57 amino acids of the Pcdh-α A cytoplasmic tail. These learning abnormalities were, however, not seen in Pcdh-αΔB/ΔB mice [in which the neomycin-resistance (neo) gene cassette was removed from the Pcdh-αΔBneo/ΔBneo alleles], in which the expression level of the Pcdh-α A isoforms was recovered, although the Pcdh-α B isoforms were still completely missing in the brain. In addition, the amount of 5-hydroxytryptamine increased in the hippocampus of the hypomorphic Pcdh-α A mutant mice but not in recovery Pcdh-αΔB/ΔB. These results suggested that the level of Pcdh-α A isoforms in the brain has an important role in regulating learning and memory functions and the amount of 5-hydroxytryptamine in the hippocampus.