We studied cell-volume changes caused by adding corticostatin (CS) or defensin-like peptides to villus enterocytes isolated in suspension from guinea pig jejunum. Guinea pig CS (10(-9) M) added to villus cells in Na(+)-containing medium reduced volume, but immediate cell swelling was caused by 10(-6) M guinea pig CS. In Na(+)-free N-methyl-D-glucamine-containing medium 10(-9) M guinea pig CS accelerated the initial rate of shrinkage compared with cells in N-methyl-D-glucamine-containing medium alone as well as causing greater cell shrinkage. Guinea pig CS-stimulated cell shrinkage was prevented by a Ca2(+)-channel blocker--5 microM nifedipine, by chelation of extracellular Ca2+ with 100 microM EGTA, or by omega-conotoxin (10(-9) M). The Ca2+ ionophore A23187 (2.5 microM) reduced volume when added to villus cells in N-methyl-D-glucamine-containing medium; this action was prevented by EGTA, or quinine--an inhibitor of K+ conductance, or 9-anthracenecarboxylic acid--a Cl- channel blocker, suggesting that the volume reduction occurred because K+ and Cl- conductances were activated. Guinea pig CS-stimulated volume reduction was also prevented by 100 microM quinine or 9-anthracenecarboxylic acid. We conclude that jejunal villus enterocytes possess a Ca2(+)-activated Cl- conductance and a K+ conductance that need not be stretch-activated. Corticostatic peptides cause volume reduction in villus cells by activating L-type Ca2+ channels; other defensin-like peptides were without effect.