Biomedical subjects
T L Valentino
Publications and source records attributed to T L Valentino.
Immunocytochemical localization of polyamines in the tiger salamander retina.
The polyamines spermine and spermidine are present in neural tissue, but their functions there are not well understood. Recent work suggests that the NMDA subtype of glutamate receptors, other glutamate receptor subtypes, and certain K(+)-channels, are neural targets for polyamines. To better understand the neuron-specific roles of polyamines, we have developed antibodies that interact with spermine and spermidine in aldehyde-fixed tissue and used these antibodies in immunocytochemical studies to determine the cellular localization of these polyamines in the tiger salamander retina. The affinity-purified, polyclonal antibodies were highly specific for spermine and spermidine, exhibiting < 1% cross reactivity with putrescine, and virtually no cross-reactivity with GABA, arginine, lysine, or glutaraldehyde. Polyamine labeling was most abundant in cells in the inner half of the inner nuclear layer and in the ganglion cell layer. Some cells in the outer half of the inner nuclear layer are labeled, and there was some labeling in both synaptic layers. Double-labeling experiments indicated (1) all GABAergic amacrine cells were polyamine-positive; and (2) all ganglion cells (identified by back-filling after microinjections of rhodamine in the optic nerve) were polyamine-positive. These results are consistent with a role for polyamines as modulators of NMDA receptor function and channel function in the inner retina.
Retinal pigment epithelial repopulation in monkeys after submacular surgery.
BACKGROUND: Transplantation of retinal pigment epithelium may be a treatment for retinal diseases, such as age-related macular degeneration and hereditary macular degeneration. Before transplantation studies are undertaken, questions concerning repopulation of retinal pigment epithelial cells in situ and photoreceptor repair after submacular surgery need to be addressed. METHODS: We removed the retinal pigment epithelium from Bruch's membrane in the macaque monkey in the macula and outside the vascular arcades. This model allowed the study of in situ retinal pigment epithelium regrowth and photoreceptor repair for 9 months following débridement. RESULTS: Fluorescein angiography revealed a window defect in the area of denuded retinal pigment epithelium. Histologic studies revealed repopulated nonpigmented retinal pigment epithelial cells in the denuded areas in both the early and late periods. At 9 months, the repopulated retinal pigment epithelium was associated with repaired, normal-appearing photoreceptor outer segments. Retinal pigment epithelium regrowth was observed only if Bruch's membrane was intact. CONCLUSIONS: Repopulation of retinal pigment epithelium in the adult primate can occur rapidly and can support the repair of damaged photoreceptors following submacular surgery.
Débridement of the pig retinal pigment epithelium in vivo.
OBJECTIVE: To study the morphologic effects of surgical débridement of the retinal pigment epithelium (RPE) in an animal model. METHODS: A pars plana vitrectomy was performed in the domestic pig, and a neurosensory retinal detachment was created by injecting the calcium-chelating agent edetic acid (commonly referred to as ethylenediaminetetraacetic acid or EDTA) into the subretinal space through a retinotomy. Twenty minutes later, the RPE was débrided by gently brushing Bruch's membrane with a soft-tip silicone catheter. Dissociated RPE was aspirated from the subretinal space, and the retina was reattached with a fluid-gas exchange. RESULTS: Light microscopic analysis confirmed that Bruch's membrane was devoid of native RPE and the choriocapillaris was morphologically intact immediately after débridement. Photoreceptor outer segments were disrupted and foreshortened immediately after RPE débridement. One to 4 weeks later, a layer of hypopigmented RPE covered most of the previously débrided areas of Bruch's membrane. The choriocapillaris was intact in areas of Bruch's membrane that were repopulated by hypopigmented RPE, and remained intact 12 weeks after débridement. Some regions of Bruch's membrane near the retinotomy remained devoid of RPE for more than 4 weeks after débridement. The choriocapillaris was atrophic and there was extensive disruption of the outer retinal layers in these areas. CONCLUSIONS: The RPE healed in most areas after surgical débridement of the RPE in the experimental animal. Atrophy of the choriocapillaris was present in areas of poor RPE healing near the retinotomy.
Desensitizing glutamate receptors shape excitatory synaptic inputs to tiger salamander retinal ganglion cells.
AMPA/kainate (KA) receptors mediate a component of ganglion cell excitatory postsynaptic currents (EPSCs). We investigated whether desensitization at these receptors contribute to the shape of transient EPSCs in ON-OFF ganglion cells. Whole-cell, voltage-clamp recordings were made from ganglion cells in the retinal slice or in isolation. EPSCs were evoked by either stimulating the slice with light or puffing K+ at the outer plexiform layer (OPL). The AMPA/KA receptor-mediated component of the EPSCs was isolated by including NMDA receptor antagonists in the bath. Strychnine and picrotoxin blocked inhibitory inputs. In isolated ganglion cells, cyclothiazide (10 microM), which blocks desensitization in non-NMDA receptors, enhanced both the amplitude and the duration of currents evoked by puffs of AMPA or glutamate. EPSCs evoked by K(+)-puffs in the OPL were also enhanced by cyclothiazide (30 microM). When AMPA/KA receptors were blocked with NBQX (10 microM), no enhancement of the EPSCs by cyclothiazide was observed, indicating that cyclothiazide did not act presynaptically. Cyclothiazide also enhanced the amplitude and duration of both the ON and OFF light-evoked (L-) EPSCs recorded in ON-OFF ganglion cells. Current-voltage relationships showed the enhancement was not voltage dependent. When control and enhanced responses where normalized, it was observed that the rate of desensitization of both the ON and OFF L-EPSCs was decreased by cyclothiazide. Cyclothiazide selectively enhanced the AMPA/KA receptor-mediated component of ganglion cells EPSCs, suggesting that desensitization of AMPA/KA receptors shape transient L-EPSCs.
Photoreceptor transplantation: anatomic, electrophysiologic, and behavioral evidence for the functional reconstruction of retinas lacking photoreceptors.
We have investigated the possibility of using transplantation of immature or mature rodent photoreceptors as well as mature human photoreceptors to reconstruct retinas in which photoreceptor degeneration is either inherited or environmentally induced. To this end, we have devised methods for isolating and transplanting the outer nuclear layer (ONL) (e.g., the photoreceptor layer) to the subretinal space of mature rodents. In addition we found that if portions of the inner retina are transplanted along with the intact photoreceptor sheet, photoreceptor organization is better maintained. In ultrastructural studies of the reconstructed retina an outer plexiform-like layer (OPL) is visible at the interface of the transplanted ONL and the host inner nuclear layer, with invaginating ribbon synapses characteristic of those formed by rod photoreceptors evident within this OPL. Ribbon synapses are found only rarely in unreconstructed retina. These results suggest that synaptic connections between transplanted photoreceptors and host cells may be made. Evidence for the potential recovery of function following photoreceptor transplantation is found in visually evoked cortical responses and behavioral responses (pupillary reflex) to light stimulation of the reconstructed eye. These findings suggest the possibility that neural transplantation can reconstruct a sensory end organ--in this case the retina--to restore evoked activity and an appropriate behavioral response to sensory stimulation.