Use of aggregating brain cultures to study the replication of herpes simplex virus types 1 and 2 in central nervous...

A novel tissue culture system consisting of reaggregated embryonic mouse brain cells was used to examine the replication of herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) within central nervous system tissue. Brain aggregates cultured 30-40 days in vitro demonstrated progressive maturation and differentiation into cells recognizable as neurons, astrocytes and oligodendrocytes, with the latter cell type exhibiting myelin production. Mature aggregates were infected with HSV and sampled at timed intervals postinfection for morphological, virological, and biochemical assays. By electron microscopy mature nucleocapsids were observed in the nucleus of peripheral cells at 9 h and in all cell types by 33 h. Virus-specific antigens were observed, using the immunoperoxidase test, within peripheral cells at 12 h postinfection (p.i.). By 24 h p.i., antigen production had progressed throughout the infected aggregates. Growth curves of HSV-1 and HSV-2 for intracellular and extracellular infectious virus production correlated well with virus-induced morphological changes and antigen production. SDS-polyacrylamide slab gel electrophoresis of isotopically-labelled proteins and glycoproteins synthesized from 4 to 24 h p.i. in virus-infected aggregates revealed typical HSV-1 polypeptide profiles and HSV-1 and HSV-2 glycoprotein profiles. Our results suggest that aggregating brain cultures may provide a useful and more accurate in vitro model for the study of HSV-induced neurological disease.