The common earthworm Aporrectodea caliginosa survives drought by forming estivation chambers in the topsoil under even very slight reductions in soil water activity. We induced estivation in a soil of a consistency that allowed the removal of intact soil estivation chambers containing a single worm. These estivation chambers were exposed to 97% relative humidity for 30 d to simulate the effect of a severe summer drought. Gas exchange, body fluid osmolality, water balance, urea, and alanine were quantified, and whole-body homogenates were screened for changes in small organic molecules via (1)H-nuclear magnetic resonance (NMR). Formation of estivation chambers was associated with a dramatic increase in body fluid osmolality, from 175 to 562 mOsm kg(-1), accompanied by a 20% increase in water content. Dehydration for 1 mo caused a further increase to 684 mOsm kg(-1), while the worms lost 50% of their water content. Gas exchange was depressed by 50% after worms entered estivation and by 80% after a further 30 d of dehydration. Urea concentrations increased from 0.3 to 1 micromol g(-1) dry mass during this time. Although (1)H-NMR did not provide the identity of the osmolytes responsible for the initial increase in osmolality after estivation, it showed that alanine increased to more than 80 mmol L(-1) in the long-term-estivation group. We propose that alanine functions as a nitrogen depot during dehydration and is not an anaerobe product in this case.