Three soluble receptor-resistant (srr) mutants, srr7, srr11 and srr18, derived from a highly neurotropic mouse hepatitis virus (MHV) JHMV have a single amino acid mutation in the spike (S) protein. We examined using ICR mice whether the amino acids mutated in the mutants were involved in the neurovirulence. Srr7 showed apparently reduced neurovirulence relative to the wild-type (wt) JHMV in terms of the LD50 and survival time, while the others showed slightly reduced virulence. In the brain and spinal cord, the growth of srr7 was more than 2 log10 lower than that of the wt virus. Histopathologically, no significant difference was revealed between wt and srr7-infected mice on day 2 postinoculation (p.i.), with only scant inflammation and a minimum degree of neuropathological changes. The major difference was that apoptotic cells were frequently encountered in the srr7-infected mouse brain, but not in wt-infected mice on day 2 p.i. However, there was no difference between these viruses in the potential to induce apoptosis in cultured cells. The apoptosis in the brain did not appear to result from the direct viral attack, since apoptotic cells were found in the lesion where viral antigens were barely detected. The present study suggests that the amino acids mutated in the S protein of srr mutants, especially the amino acid at position 1114 mutated in srr7, influence the neurovirulence in mice.