The cecum of the germ-free rat is filled with a large volume of liquid: cecum plus cecal contents comprise up to 25% of the rodent's body weight. To explain the presence of the liquid cecal contents in the germ-free rat, cecal transport of water and electrolytes was studied using a closed-loop technique with [14C]polyethylene glycol as a nonabsorbable marker. When NaCl (154 mEq/liter) was instilled into germ-free rat ceca, absorption of water, Na, and Cl occurred and was similar to that in conventional animals. In contrast, when an equal volume of supernatant from germ-free cecal contents was instilled into germ-free rat ceca, secretion of water, Na, and Cl occurred. Similarly, when the germ-free supernatant was instilled into ceca of conventional rats, secretion of water, Na, and Cl occurred at rates equal to that seen in the germ-free rat. Therefore, it appeared that the composition of the germ-free cecal contents was primarily responsible for the cecal secretion. Analysis of the germ-free cecal contents to determine what caused the cecal secretion revealed very low measurable anions (Cl<2, HCO3<2 mEq/liter), a low Na concentration, slight hyperosmolality as determined by freezing-point depression, and elevated colloid osmotic pressure. Fluid made up with Na2SO4 to resemble the germ-free cecal supernatant in ionic composition and absence of exchangeable anions produced cecal secretion equal to that of the germ-free rat cecal contents. Fluid simulated to reproduce the effects of the low Na concentration and also the colligative properties and colloid osmotic pressure had minimal effects on cecal transport. These studies demonstrate that cecal enlargement in the germ-free rat is associated with cecal secretion of water and electrolytes, that germ-free ceca can transport water and electrolytes in a normal fashion, and that cecal secretion in the germ-free rat is due primarily to the absence of permeable anions in the germ-free cecal contents.