Establishment of a quiescent herpes simplex virus type 1 infection in neurally-differentiated PC12 cells

Rat pheochromocytoma (PC12) cells differentiated with nerve growth factor ( Nd-PC12) were used to investigate the establishment of a non-productive her pes simplex virus type 1 (HSV-1) infection that is reversible. The results of this work are as follows: (i) Nd-PC12 cultures could be maintained as lo ng term (>7 weeks) non-dividing cultures only when plated on collagen-coate d dishes in the absence of serum; (ii) Infection of Nd-PC12 with HSV-1 stra ins KOS and 17 in the transient presence of acycloguanosine (ACV) resulted in all cultures free of detectable levels of infectious virus at the time o f ACV removal and ACV was not needed to maintain the non-productive quiesce nt state in the subsequent 8 weeks; (iii) These persistently infected and q uiescent (QIF)-PC12 cultures demonstrated both spontaneous and forskolin-in ducible virus production, at low (5%) and high frequencies (92 - 100%), res pectively during the first 2 weeks post-ACV withdrawal. (iv) In contrast to other in vitro models, HSV-1 failed to reactivate following removal of ner ve growth factor. (v) A high percentage of QIF-PC12 cultures (50-100%) prod uced virus in response to forskolin treatment as long as 7 weeks post-ACV w ithdrawal. (vi) Expression of HSV-1 productive genes (i.e. alpha 0, alpha 4 , alpha 27, U-L,30 and U-L,18) dropped precipitously in the presence of ACV and remained undetectable or continued to decline following its removal, w hereas the levels of LAT and the host gene G3PDH remained relatively consta nt throughout the 31 day study period as measured by RT-PCR. These results indicate that QIF-PC12 cells offer a novel, neuronal cell culture system th at may enhance our ability to study HSV-1 reactivation from a cryptic, late nt-like, non-productive state in the absence of replication inhibitors.