The effects of HI-6 and pralidoxime chloride (2-PAM) on soman-induced lethality, time to death and several cholinergic parameters in rats were compared to understand the beneficial action of HI-6. Treatment with atropine sulfate (ATS) or HI-6 alone protected against 1.2 and 2.5 LD50s of soman respectively, whereas 2-PAM or methylated atropine (AMN) alone afforded no protection. Addition of ATS, but not AMN, to HI-6-treated rats enhanced the protection from 2.5 to 5.5 LD50s. HI-6 increased the time-to-death, while 2-PAM had no effect; a combination of HI-6 and ATS provided the most significant increase in time-to-death. Cholinesterase (ChE) activity was not altered in any tissue by ATS, HI-6 or 2-PAM treatment individually, but was markedly inhibited in all tissues by 100 micrograms/kg of soman. In soman-poisoned rats, the HI-6, but not the 2-PAM, group had significantly higher levels of ChE in blood and other peripheral tissues than did the group given soman alone. Neither HI-6 nor 2-PAM affected soman-inhibited ChE in the brain. Additional ATS treatment had no effect on ChE activity. HI-6 and 2-PAM neither modified baseline brain acetylcholine (ACh) or choline (Ch) levels nor protected against soman-induced ACh or Ch elevation. 2-PAM exhibited a 4-fold more potent in vitro inhibition of 3H-quinuclidinyl benzilate (3H-QNB) binding and sodium-dependent high-affinity Ch uptake (HACU) than did HI-6 in brain tissues. The findings that 2-PAM is a more potent in vitro inhibitor of muscarinic receptor binding and HACU than HI-6, and yet neither elevates ChE activity in the periphery nor protects rats against soman poisoning, indicate the importance of higher ChE activity in the periphery of HI-6-treated rats. Maintenance by HI-6 of a certain amount of active ChE in the periphery appears to be important for survival after soman exposure.