The role of herpes simplex thymidine kinase expression in neurovirulence and latency in newborn vs. adult mice.

Infection by standard thymidine kinase-positive (TK+) and TK- mutant herpes simplex virus (HSV) was performed in order to evaluate the role of HSV TK expression in neurovirulence and in HSV latency. In newborn mice, mortality and trigeminal ganglion (TG) HSV titer correlated (both were high) for TK+ and TK- HSV. In adult mice after TK- HSV infection they also correlated (both were low). After TK+ infection of adult mice, correlation was not present; mortality was low while HSV titer was moderately high. During the period of HSV latent infection (> 28 days after HSV infection), the number of neurons expressing HSV latency-associated transcript (LAT) was much greater for TK- HSV newborn-inoculated mice (average of 943/ganglion) than adult-inoculated mice (average of 138/ganglion). In addition, total amount of TG LAT was greater in the former than the latter. Reactivation from latency was restricted, however, for both groups. This result supported the important role of HSV TK expression in HSV reactivation, even when the number of LAT-positive neurons was greatly increased. The following conclusions were drawn from the study of TK- HSV in newborn mice: (i) HSV TK expression was of limited importance for neurovirulence and in vivo HSV TG infection (but was of importance in adult mice); (ii) increased in vivo HSV TG infection correlated with increased number of LAT-positive neurons, so that HSV replication and establishment of latency were not completely separable; and (iii) even with greatly increased numbers of latently infected neurons, HSV TK expression was important for reactivation from latency. Results in newborn mice suggested that the role of HSV TK expression in reactivation from latency and in neurovirulence were separable. To further investigate HSV replication in newborn and adult mice, ganglia were infected with HSV in vitro and either maintained in vitro or transplanted beneath the renal capsule of adult recipients. In both of these studies, HSV titers in ganglia were much higher in newborn than adult ganglia. This suggested that in addition to the well-know role of the immune system in HSV neurovirulence in newborn mice, it is likely that HSV replication per se in neural tissue is greater in newborn than adult mice. This may be related to the high level of HSV neurovirulence in newborn mice.

Immature and mature neurofilament-immunoreactive trigeminal fibers can innervate the iris as studied by intraocular grafting of iris and trigeminal ganglia.

Using immunohistochemistry on stretch-prepared whole mounts of adult rat irides, a dense, well-organized plexus of neurofilament-positive nerves originating in the trigeminal ganglion can be visualized. Such a two-dimensional tissue preparation is well-suited for studies on sensory and autonomic nerve fiber growth. In the present study the growth capacity of such neurofilament-positive nerves has been studied immunohistochemically. In irides homologously transplanted to the anterior eye chamber of adult albino rats, the intrinsic neurofilament-positive network had almost completely disappeared 4 days postoperatively. In whole mounts of iris grafts after 15 days and 4 weeks in oculo a gradually increasing plexus of nerves was observed. After 3.5 months in oculo a dense, regular network of fluorescent fibers had formed in the iris grafts to the same magnitude as in situ. However, whereas large axon bundles constituted a prominent feature of the distribution of neurofilament-positive nerves in situ, only a few and relatively thin axon bundles were seen in the grafts. The growth capacity of the neurofilament-positive trigeminal nerves was also studied by grafting fetal trigeminal ganglia to the anterior eye chamber. As visualized in cryostat sections, trigeminal grafts contained a large number of strongly fluorescent perikarya and a high density of positive fibers after intraocular maturation. Such grafts readily innervated the host iris. In the area immediately adjacent to the grafts, thin, parallel, rather weakly fluorescent fibers radiated out from the ganglia. When mature trigeminal grafts with attached host iris were regrafted to the anterior eye chamber of adult animals for a few days, in order to remove the intrinsic host iris innervation, such irides showed outgrowing fibers, often organized in small axon bundles, at long distances from the ganglion graft. The present report shows that both mature and immature neurofilament-immunoreactive neurons are capable of innervating the iris. Furthermore, this ingrowth can occur both in the presence and absence of normal intrinsic neurofilament-positive nerve fibers.