Fractional lithium clearance (CLi/CIn), transit time-occlusion time (e-TT/OT), and late proximal tubular fluid-to-plasma inulin ratio [1/(TF/P)In] collected by micropuncture were determined successively in the same rat during Amytal anesthesia. The rats were examined during hydropenia, after partial aortic constriction, or during saline diuresis. There was a linear relationship (r = 0.80) between CLi/CIn and e-TT/OT. The 1/(TF/P)In ratio correlated closely with both CLi/CIn (r = 0.88) and e-TT/OT (r = 0.91) when intraluminal pressure was maintained at the free-flow level during fluid collection. If fluid collection was guided merely by the position of an oil droplet and the luminal diameter, the 1/(TF/P)In data were not correlated with either CLi/CIn or e-TT/OT. Over a wide range of proximal absolute and fractional reabsorption rates the technically simpler lithium clearance and TT/OT methods may provide data on proximal fractional reabsorption that are as accurate and reliable as data obtained by pressure-controlled micropuncture collection. Micropuncture carried out without pressure control provides highly inaccurate data and is clearly inferior to the other methods. These results are consistent with the possibility that lithium is reabsorbed exclusively by the proximal tubules, 17-20% being reabsorbed by the pars recta.