A Naturally Occurring Canine Model of Autosomal Recessive Congenital Stationary Night Blindness

Congenital stationary night blindness (CSNB) is a non-progressive, clinically and genetically heterogeneous disease of impaired night vision. We report a naturally-occurring, stationary, autosomal recessive phenotype in beagle dogs with normal daylight vision but absent night vision. Affected dogs had normal retinas on clinical examination, but showed no detectable rod responses. They had “negative-type” mixed rod and cone responses in full-field ERGs. Their photopic long-flash ERGs had normal OFF-responses associated with severely reduced ON-responses. The phenotype is similar to the Schubert-Bornschein form of complete CSNB in humans. Homozygosity mapping ruled out most known CSNB candidates as well as CACNA2D4 and GNB3. Three remaining genes were excluded based on sequencing the open reading frame and intron-exon boundaries (RHO, NYX), causal to a different form of CSNB (RHO) or X-chromosome (NYX, CACNA1F) location. Among the genes expressed in the photoreceptors and their synaptic terminals, and mGluR6 cascade and modulators, reduced expression of GNAT1, CACNA2D4 and NYX was observed by qRT-PCR in both carrier (n = 2) and affected (n = 2) retinas whereas CACNA1F was down-regulated only in the affecteds. Retinal morphology revealed normal cellular layers and structure, and electron microscopy showed normal rod spherules and synaptic ribbons. No difference from normal was observed by immunohistochemistry (IHC) for antibodies labeling rods, cones and their presynaptic terminals. None of the retinas showed any sign of stress. Selected proteins of mGluR6 cascade and its modulators were examined by IHC and showed that PKCα weakly labeled the rod bipolar somata in the affected, but intensely labeled axonal terminals that appeared thickened and irregular. Dendritic terminals of ON-bipolar cells showed increased Goα labeling. Both PKCα and Goα labeled the more prominent bipolar dendrites that extended into the OPL in affected but not normal retinas. Interestingly, RGS11 showed no labeling in the affected retina. Our results indicate involvement of a yet unknown gene in this canine model of complete CSNB.     

Developmental expression and intracellular location of P400 protein characteristic of Purkinje cells in the mouse cerebellum

The developmental expression and intracellular localization of a cerebellum-characteristic 250-kDa glycoprotein, P400 protein, were studied by immunohistochemical and immunoblot methods using a monoclonal antibody against P400 protein. In the cerebellum of normal mouse, the expression of P400 protein increased from Postnatal Day 3 to Day 21. This enhancement of P400 protein expression occurred only in the Purkinje cells and proceeded with the growth of their dendritic arborization. Electron microscopic analysis indicated that P400 protein is present at the plasma membrane, the endoplasmic reticulum, and the postsynaptic densities of Purkinje cells. Immunohistochemistry of the cerebella of neurological mutant mice indicated that the Purkinje cells of reeler, weaver, and pcd mutant mice retain the ability to produce a large amount of P400 protein. However, the Purkinje cells of staggerer mutant mouse proved to be incapable of enhanced P400 protein expression. These results indicate that P400 protein is a Purkinje cell-characteristic plasma membrane-associated glycoprotein, which is also present at the postsynaptic density and endoplasmic reticulum and that the expression of P400 protein in Purkinje cells is closely associated with the growth of their dendritic arborization.     

Starvation-induced changes in rat brain corticotropin-releasing factor (CRF) and pituitary-adrenocortical response

Starvation-induced changes in CRF concentration in major brain regions and abnormalities in the pituitary-adrenal axis were examined in rats using rat CRF radioimmunoassay. The CRF concentrations in the hypothalamus and cerebellum were significantly reduced in the completely starved rats, while those in the midbrain, thalamus and neurointermediate lobe of the pituitary were significantly increased in the semi-starved or completely starved rats. No significant changes in the CRF concentrations were found in the pons, medulla oblongata and cerebral cortex. In the completely starved rats, the serum ACTH level was significantly reduced, whereas the serum corticosterone level was markedly elevated. These observations suggest that starvation may stimulate the CRF-ACTH-corticosterone system and that not only hypothalamic CRF but also extrahypothalamic CRF may be discretely related to feeding behavior or starvation. The reduced serum ACTH level in starved rats may be ascribed to the negative feedback effect of the elevated serum corticosterone.     

Synthesis and characterization of intermediate and transition-state analogue inhibitors of gamma-glutamyl peptide ligases

The phosphonodifluoromethyl ketone and phosphonofluoridate derivatives of L-glutamic acid were synthesized and characterized as analogues of the gamma-glutamyl phosphate intermediate and the tetrahedral transition state, respectively, for the inhibition of gamma-glutamylcysteine synthetase and glutamine synthetase. The former served as a poor inhibitor of both enzymes, but the latter inhibited glutamine synthetase with a Ki of 59 microM and partially inactivated the enzyme in an NH3- and ATP-dependent manner.     

Expression of foreign type I ribulose-1,5-bisphosphate carboxylase/ oxygenase (EC 4.1.1.39) stimulates photosynthesis in cyanobacterium Synechococcus PCC7942 cells

A reporter gene assay revealed that promoters derived from Synechococcus PCC7942 (S.7942) psbAI and Synechocystis PCC6803 (S.6803) psbAII were suitable for the expression of foreign ribulose-bisphosphate carboxylase (RuBisCO; EC 4.1.1.39) in S.7942 cells. Transformational vectors with a promoter and a foreign RuBisCO gene, cvrbc originated from Allochromatium vinosum, were constructed on a binary vector, pUC303, and introduced to S.7942 cells. When the cvrbc was expressed with the S.7942 psbAI promoter, the total RuBisCO activity increased 2.5- to 4-fold than that of the wild type cell. The S.6803 psbAII promoter increased the activity of the transformant 1.5–2 times of that of wild type cell. There was a significant increase in the rate of photosynthesis depending on the increase of RuBisCO activity. The maximum rate of photosynthesis of the transformant cell was 1.63 times higher than that of the wild type under the illumination of 400 μmol m⁻² s⁻¹, at 20 mM bicarbonate and at 30 °C. Although the photosynthesis of the higher plant is limited by the ability of photosystems under high irradiance and the high CO₂ concentration, that of the S.7942 cell is limited by the RuBisCO activity, even at high CO₂ concentrations and under high irradiance.     

Simultaneous spawning by female stream goby Rhinogobius sp. and the association with brood cannibalism by nesting males

A laboratory experiment was conducted by varying the undersurface area of nesting substratum and the number of females in an experimental tank to elucidate the determinants of the mating pattern in the stream goby, Rhinogobius sp. cross‐band type. Males with larger nests tended to attract two or more females to their nest in a tank. Moreover, males spawned simultaneously with multiple females and entire brood cannibalism by males was rarely observed under a female‐biased sex ratio. When males spawned with a single female with low fecundity, however, entire brood cannibalism occurred at a high frequency, suggesting that a male guarding a nest with fewer eggs consumes the brood. Therefore, spawning behaviour of females that leads to a large egg mass would decrease the risk of entire brood cannibalism. In this species, simultaneous spawning by multiple females in a nest serves as a female counter‐measure against entire brood cannibalism. These results suggest that a conflict of interest between the sexes through brood cannibalism is a major determinant of simultaneous spawning.     

An anticomplementary agent, K-76 monocarboxylic acid: its site and mechanism of inhibition of the complement activation cascade.

A monocarboxylic acid derivative (K-76 COOH) of K-76, purified from the culture filtrate of Stachybotrys complement I nov. sp. K-76, inhibits complement (C) activity. Its inhibitory action is mainly on C5 step. It strongly inhibits the generation of EAC1,4b,2a,3b,5b from C5 and EAC1,4b,2a,3b, and accelerates the decay of EAC1,4b,2a,3b,5b. It also causes some inhibition of the reactions of the reactions of C2,C3,C6,C7 and C9 with their respective preceding intermediate cells. It has no effect on the generation of EAC1,4b from C4 and EAC1, or of EAC-8 from C8 and EAC-7, and apparently increases the generation of EAC1,4b from C1 and EAC4b probably by inhibiting transfer or turnover of C1. It does not affect the rate of decay of EAC1,4b,2a or the T max of generation of EAC1,4b,2a, and it inhibits immune adherence only at high concentration. K-76 COOH also strongly inhibits hemolysis through the alternative pathway of C activation by cobra venom factor, but it does not seem to inhibit the early steps of the alternative pathway, because it has little affect on the consumption of C3 or the conversion of beta 1C to beta 1A on treatment of C serum with zymosan. K-76 COOH probably combines with C5 molecules, forming the inactive complexes, or it causes the structural alteration of C5.     

Structural factors contributing to the Abl/Lyn dual inhibitory activity of 3-substituted benzamide derivatives

To investigate why 3-substituted benzamide derivatives show dual inhibition of Abl and Lyn protein tyrosine kinases, we determined their inhibitory activities against Abl and Lyn, carried out molecular modeling, and conducted a structure-activity relationship study with the aid of a newly determined X-ray structure of the Abl/Lyn dual inhibitor INNO-406 (formerly known as NS-187) bound to human Abl. We found that this series of compounds interacted with both kinases in very similar ways, so that they can inhibit both kinases effectively.