Cellular and subcellular localization of PDE10A, a striatum-enriched phosphodiesterase.

PDE10A is a recently identified phosphodiesterase that is highly expressed by the GABAergic medium spiny projection neurons of the mammalian striatum. Inhibition of PDE10A results in striatal activation and behavioral suppression, suggesting that PDE10A inhibitors represent a novel class of antipsychotic agents. In the present studies we further elucidate the localization of this enzyme in striatum of rat and cynomolgus monkey. We find by confocal microscopy that PDE10A-like immunoreactivity is excluded from each class of striatal interneuron. Thus, the enzyme is restricted to the medium spiny neurons. Subcellular fractionation indicates that PDE10A is primarily membrane bound. The protein is present in the synaptosomal fraction but is separated from the postsynaptic density upon solubilization with 0.4% Triton X-100. Immuno-electron microscopy of striatum confirms that PDE10A is most often associated with membranes in dendrites and spines. Immuno-gold particles are observed on the edge of the postsynaptic density but not within this structure. Our studies indicate that PDE10A is associated with post-synaptic membranes of the medium spiny neurons, suggesting that the specialized compartmentation of PDE10A enables the regulation of intracellular signaling from glutamatergic and dopaminergic inputs to these neurons.

A poorly differentiated germ cell tumor (seminoma) in a Long Evans rat.

A large neoplasm that replaced 1 testis of a Long Evans Rat was noted at the final necropsy of a dietary 2-yr study. By light microscopy, the morphological features were consistent with a poorly differentiated seminoma. Ultrastructurally, the cells were polygonal, had a round nucleus, had straight cellular boundaries, and bore no resemblance to Sertoli cells. Although there was little evidence of spermatocytic differentiation, the presence of proacrosomal granules and vesicles, prominent Golgi apparatus, tight intercellular junctions, and a few centriolar pairs without axoneme development, in conjunction with the absence of lipid droplets or abundant smooth endoplasmic reticulum, supported the diagnosis of seminoma rather than Leydig cell tumor. The cells were S-100- and vimentin-positive, although cytokeratin- and alpha-fetoprotein-negative. Seminomas are extremely rare neoplasms in rats; this is the first report in this strain and the first extensive analysis of a rat seminoma without spermatocytic differentiation.

Changes in rat heart histomorphometry due to two-week dietary restriction.

In preclinical safety studies in which the administration of a test compound causes reductions in food consumption, body weights, and organ weights, it may be difficult to differentiate direct compound-induced effects on organ weights from those simply due to reduced nutrition. To address this problem in reference to the heart, hearts were obtained from rats that were known to have had reductions in body weights and absolute heart weights as a result of feed restriction. Rats (40/sex) were divided into 4 groups (10/sex) and given quantities of ad libitum diet for 2 wk as follows: Group 1, 100%; Group 2, 75%; Group 3, 50%; and Group 4, 25%. Routine histologic evaluation was performed on longitudinal sections of paraffin-embedded hearts stained with hematoxylin and eosin. Computer-assisted image analysis was conducted on left ventricular free-wall of picrosirius red-stained sections for histomorphometric evaluation of ratio of cross-sectional area occupied by myofibers versus interstitium and for quantification of myofiber width. No differences were detected histologically among groups, and no difference in the mean myofiber: interstitium ratio was detected between Groups 1 (9.1) and 4 (9.5). Mean values for myofiber width ranged from 24.6 microns for Group 1 to 17.3 microns for Group 4. Two-way ANOVA revealed a strong effect of dietary restriction on reduction of myofiber width but no consistent gender effect. The significant dietary effects occurred in Groups 3 and 4 compared to corresponding controls. The present authors speculate that, if reductions in feed intake were < 50% in short-term preclinical studies, any reductions in myofiber width could imply a primary test article effect. Conversely, if reductions in feed intake were > or = 50% in such studies, reductions in myofiber width could be caused either solely by inadequate nutrition or by combined effects of nutrition and test article.

Tapetal effect of an azalide antibiotic following oral administration in beagle dogs.

An azalide antibiotic (CP-62,993) was administered at 100 mg/kg by oral gavage once daily for 35 consecutive days to 3 normal Beagle dogs (tapetal) and 3 Beagle dogs lacking a clinically apparent ocular tapetum (atapetal). The total dose delivered was approximately 100-fold the recommended clinical dose. Bilateral ophthalmoscopic changes were observed in the treated tapetal dogs on Day 36, consisting of mild to moderate tapetal decoloration with loss of the normal color change at the junction with the nontapetal fundus and muting of reflectivity of the normally highly reflective tapetum; treated atapetal and all control tapetal and atapetal dogs had no ophthalmoscopic changes. Microscopic examination of ocular tissue revealed rudimentary tapetal cell layers in the correct location in untreated, clinically atapetal eyes. Tapetal cells from treated tapetal and atapetal dogs were swollen and vacuolated, and contained intracytoplasmic, electron-dense debris but no recognizable tapetal rodlets. Lysosomal lamellar bodies were observed in the retinal ganglion cells of both treated groups and were neither enhanced nor reduced by the presence of a functional tapetum. Necrosis and inflammation were not observed in any ocular tissue. The altered ophthalmoscopic appearance of treated tapetal dogs was not influenced by the retinal changes because any effect on retinal transparency would have been seen in treated atapetal dogs. The decoloration and muting of reflectivity observed clinically in the tapetal fundus of dogs following prolonged exposure to high levels of CP-62,993 result from unique changes within the ocular tapetum itself and cannot be interpreted to be of consequence to nontapetal species including humans.