Ichiro Ishimoto

Marked increase in glutamate-aspartate transporter (GLAST/GluT-1) mRNA following transient retinal ischemia

We have demonstrated the cellular localization of glutamate-aspartate transporter (GLAST/GluT-1) mRNA in the rat retina and its induction after ischemia by in situ hybridization. GLAST mRNA was expressed in the inner two-thirds of the inner nuclear layer (INL) and in sparse small cells in the inner portion of the ganglion cell layer (GCL) of the adult rat retina. GLAST mRNA was also found in about 90% of cells in the optic nerve head where more than 90% of cells express glial fibrillary acidic protein (GFAP) mRNA. Moreover, experimental occlusion of the central retinal artery followed by reperfusion for 48 h resulted in degeneration of neurons and a marked increase in GLAST mRNA expression in the INL. These findings suggest that GLAST may be expressed in Müller cells and astrocytes in the retina, and may play an important role in regulation of extracellular glutamate concentration especially under ischemic conditions.

Aldose reductase inhibitor (CT-112) eyedrops for diabetic corneal epitheliopathy.

We treated two diabetic patients with corneal epithelial disorder that resisted conventional medical therapy with topical CT-112 (5-[3-ethoxy-4-pentyloxyphenyl]-2,4-thiazolidinedione), a newly synthesized aldose reductase inhibitor. One patient had developed recurrent corneal erosion after vitrectomy and the other had spontaneously developed superficial punctate keratopathy. The corneal lesion in each patient responded to topical CT-112 in two to four weeks and was almost cleared within two months. A similar corneal lesion recurred in both patients soon after CT-112 was discontinued, but it disappeared again when the drug was resumed.

Ontogeny of neurotensin-containing neuron system of the rat: Immunohistochemical analysis—II. Lower brain stem

The ontogeny of the neurotensin neuron system in the lower brain stem of the rat was investigated by means of indirect immunofluorescent method. Neurotensin-like immunoreactivity-containing cells first appear in the primordium of the n. tractus solitarii, n. tractus spinalis nervi trigemini, reticular formation just medial to the latter nucleus, n. reticularis parvocellularis, n. laterodorsalis tegmenti, and midbrain reticular formation of the fetus at gestational day 17. At gestional day 18, neurotensin-immunoreactive cells newly appear in the n. raphe dorsalis. Between gestational day 19 and postnatal day 7, the animals show a remarkable increase in number of immunoreactive cells and fibers in various lower brain stem areas except for n. tractus spinalis nervi trigemini and n. tractus solitarii. Moreover, during this stage, neurotensin-immunoreactive cells located in the n. prepositus hypoglossi and n. vestibularis lateralis appear for the first time at birth and postnatal day 5, respectively. Since postnatal day 7, although the majority of immunoreactive cells located in the lower brain stem decrease in number as the rats grow, immunoreactive cells in the n. tractus spinalis nervi trigemini, on the contrary, increase in number from after birth until postnatal day 10, and maintain more or less their immunoreactivity even in the adult rat. In addition, neurotensin-immunoreactive cells in the nucleus of the solitary tract increase in number during the fetal period, reach the maximum content at postnatal day 7-10, and maintain their immunoreactivity even in the adult rats. Thus, the present study demonstrated that neurotensin-like immunoreactive structures appear at a very early ontogenetical stage, suggesting that neurotensin plays an important role in the development of the lower brain stem of the rat. In addition, the present study further showed that neurotensin-immuno-reactivity shows various fluctuations during the ontogeny, suggesting multiple functions of neurotensin in the central nervous system.

Localization of vasoactive intestinal polypeptide and neurotensin immunoreactivities in the avian retina

The distribution of vasoactive intestinal polypeptide (VIP)- and neurotensin (NT)-like immunoreactivities in the chicken retina was investigated by immunohistochemistry. VIP- and NT-positive cells were found throughout the chicken retina. The majority of them were located in the proximal portion of the inner nuclear layer and the processes from these cells directed to the inner plexiform layer where they ramified, suggesting that VIP- and NT-positive cells located in this region probably are amacrine cells.

Isolation of a cDNA encoding a photoreceptor cell-specific actin-bundling protein: retinal fascin

We have isolated a novel retina‐specific gene, retinal fascin, encoding a new member of actin‐bundling protein gene family, from a bovine retina cDNA library. The cDNA encodes a 492 amino acid protein which shows 36–57% amino acid identity with three vertebrate fascins, echinoid fascin and Drosophila singed gene. Northern blot analysis revealed that retinal fascin mRNA was exclusively expressed in the eye and not seen in other tissues examined. In situ hybridization analysis indicated that retinal fascin mRNA signals were found only in the inner segment of the photoreceptor layer and outer nuclear layer, indicating that retinal fascin was specifically expressed in photoreceptor cells. As fascins are actin‐bundling proteins important for constructing several intracellular structures, retinal fascin might play a pivotal role in photoreceptor cell‐specific events, such as disk morphogenesis.

Demonstration of a substance P-like immunoreactivity in retinal cells of the rat.

The distribution of substance P (SP)-like immunoreactivity in the rat retina was investigated by immunohistochemistry. SP-positive cells were found throughout the retina. The majority of them were located in the proximal portion of the inner nuclear layer and the processes from these cells directed to the inner plexiform layer where they ramified, suggesting that SP-positive cells located in this region probably are amacrine cells. Few SP-positive cells were seen within the ganglion cell layer. They were considered displaced amacrine cells.

Expression of c-fos and c-jun mRNA Following Transient Retinal Ischemia: An Approach Using Ligation of the Retinal Central Artery in the Rat

The expression of the proto-oncogenes c-fos and c-jun was examined by in situ hybridization at various timepoints following transient retinal ischemia by means of ligation of the retinal central artery in the rat. Ischemia of 90-minute duration resulted in the degeneration of neurons in both the ganglion cell layer and the inner nuclear layer at 48 hours after reperfusion. The expression of c-fos and c-jun messenger RNA throughout the entire inner nuclear layer was transiently coinduced following 90-minute retinal ischemia with a peak at 1 hour after reperfusion. This expression was also found in the ganglion cell layer at 3 hours after reperfusion. Weak signals for c-fos and c-jun mRNA were observed at 24 hours after reperfusion and returned to near control levels by 48 hours. c-jun protein expression was detected in the ganglion cell layer, the middle of the inner nuclear layer, and optic nerve head at 3 hours, but not 1 hour, after lethal ischemia/reperfusion; however, c-fos protein expression was not detected after reperfusion. Whereas no neuronal degenerative changes were found at 7 days after 30-minute ischemic retina, c-fos and c-jun messenger RNA were also induced at 1 hour postreperfusion. To our knowledge, this study is the first report to show expression patterns of immediate-early genes after retinal ischemia/reperfusion. These results suggest that changes in expression of c-fos and c-jun after transient retinal ischemia are similar to those after transient brain ischemia, and the selective occlusion of the central retinal artery will provide a useful model for studying ischemic neuronal degeneration in vivo in the rat retina.

Differentiating full thickness macular holes from impending macular holes and macular pseudoholes

AIMS The reliability of scanning laser ophthalmoscope (SLO) microperimetry in differentiating full thickness macular holes from macular pseudoholes and impending macular holes was evaluated. METHODS 106 eyes with the clinical diagnosis of full thickness macular holes, macular pseudoholes, and impending (stage 1) macular holes were examined for the presence of deep or relative scotoma using SLO microperimetry. The relation between these scotomas and the clinical diagnosis was studied. RESULTS Deep and relative scotomas were detected in all 57 eyes with clinically defined full thickness macular holes. In contrast, among 49 eyes diagnosed with macular pseudoholes or impending macular holes, no deep and only one relative scotoma was observed. The sensitivity of the presence of a deep scotoma as an indicator of the clinical diagnosis of a full thickness macular hole was 100% (57 of 57), and the specificity was 100% (49 of 49). The sensitivity of the presence of a relative scotoma was 100% (57 of 57) and the specificity was 98.0% (48 of 49). CONCLUSION With SLO microperimetry, full thickness macular holes can be precisely and objectively distinguished from other conditions that mimic macular holes.

Ontogeny of substance p-containing structures in the ocular tissue of the rat: An immunohistochemical analysis

The ontogeny of the substance P-like immunoreactivity (SPI) containing system in the ocular tissue of the rat was examined by means of the indirect immunofluorescence method. SPI-containing amacrine cells first appeared at postnatal day 4 and displaced SPI-containing amacrine cells at postnatal day 5. After this time, they developed markedly reaching their maximum content and distribution at postnatal day 14. On the other hand, SPI-containing cells in the trigeminal ganglion were first seen at gestational day 17 and reached their maximum content at birth. SPI-containing fibers in the cornea and uvea were first observed at gestational days 17-19. The SPI-containing fibers in the iris reached their maximum content at birth, while those in the cornea, choroid and ciliary body were fully developed at postnatal day 3. In the adult rats, numerous SPI structures were still seen in the ocular tissue, retina, cornea and uvea. These findings suggest that SPI might play some role in the developing ocular tissue in addition to its neurotransmitter or neuromodulator role in the adult, because SPI structures appear in the retina before establishment of synaptogenesis and in the cornea and uvea during the fetal stage.

Differential distribution of mRNA encoding cAMP-specific phosphodiesterase isoforms in the rat brain

We studied the distributions of four different cyclic AMP-specific phosphodiesterase isoform mRNAs (APDE1-4) and compared them with that of 63 kDa calmodulin-stimulated phosphodiesterase (CPDE) in the rat brain by in situ hybridization histochemistry using specific radiolabeled oligonucleotides. The distribution patterns were unique for all the APDE isoforms examined here. Although no significant signals for APDE1 could be detected anywhere in the rat brain, all other isoforms were expressed ubiquitously but unevenly and showed overlapping distribution patterns. Among all the APDE isoforms studied here, APDE3 showed the strongest and the most extensive expression. Its distribution pattern implies that it may modulate different cellular processes associated with learning and memory. Compared to APDE3, the levels of expression of APDE2 and APDE4 were weaker, the latter showing the weakest expression. Our study suggests that different isoforms of APDE are expressed together in the same class of neurons implying complex interactions among different signaling pathways, thereby mediating distinct and specific functions.