MHC class I‐restricted killing of neurons by virus‐specific CD8+ T lymphocytes is effected through the Fas/FasL, but not the perforin pathway

Induction of MHC class I genes in neurons of the central nervous system requires signals by pro‐inflammatory cytokines, in particular IFN‐γ, and the blockade of electric activity, which is known to suppress induction of MHC related genes in a highly ordered, but unusual fashion 1 , 2 . The present experiments explore the immunological function of neuronal MHC class I antigens expressed under permissive conditions. MHC class I proteins were induced in electrically silenced murine hippocampal neurons by treatment with the sodium channel blocker tetrodotoxin and recombinant IFN‐γ, conditions which also resulted in the induction of Fas molecules. The MHC class I positive neurons were challenged with CD8+ cytotoxic T lymphocytes (CTL) specific for the H2‐Db binding peptide GP33, a dominant epitope of the lymphocytic choriomeningitis virus envelope glycoprotein, or with alloreactive CTL. Single primed neurons, attacked by GP33‐specific CTL, were continuously monitored for changes in intracellular calcium ([Ca2+]i), an indicator of cytotoxic damage. MHC class I‐induced neurons pulsed with the GP33 peptide, but not a control peptide, showed a gradual and sustained increase in [Ca2+]i within 3 h following attack by GP33‐specific CTL, while in astrocytes [Ca2+]i elevation was rapid. The slow course of the neuronal response was consistent with a delayed apoptotic killing mechanism rather than rapid granule‐mediated plasma membrane lysis. Indeed, the attacked neurons bound annexin V, indicating membrane alterations preceding apoptotic cell death. In further support of apoptotic cell death, this sustained increase of [Ca2+]i levels was also observed following attack by perforin‐deficient CTL, but was not detected in neurons derived from mutant lpr mice, which lack functional Fas molecules.