Distribution of Ankyrin Isoforms and Their Proteolysis After Ischemia and Reperfusion in Rat Brain

Abstract: The distribution of brain-type ankyrin (ankyrin_B, 212 kDa) and erythrocyte-type ankyrin (ankyrin_R, 239 kDa) was investigated in the subcellular fractions of rat forebrain (P1, 1,000 g pellet; P2, 15,000 g pellet; P3, 100,000 g pellet; S, 100,000 g supernatant) by immunoblotting using specific antibodies. The P2 fraction contained ∼40% of the 212- and 163-kDa isoforms of ankyrin_B and the 239-kDa isoform of ankyrin_R. Further subfractionation of the P2 by Percoll gradient centrifugation followed by separation of myelin showed association of the three ankyrin isoforms with the synaptosome-rich fraction but not with the myelin-rich fraction. The plasma membrane-rich P3 fraction contained a concentration of ankyrin isoforms similar to that in the P2 fraction. In vitro proteolysis of ankyrin in the P2 fraction with calpain showed that the 212-kDa ankyrin_B was more susceptible to calpain than was ankyrin_R. In the two-vessel occlusion model, ischemia for 30 min generated the 160-kDa fragment of ankyrin_R, and reperfusion for 60 min after 30 min of ischemia remarkably increased the 160-kDa fragment. The reperfusion also significantly decreased the 212-kDa isoform of ankyrin_B. Both ischemia-reperfusion and in vitro proteolysis with calpain generated the 160-kDa fragment of ankyrin_R, suggesting the involvement of calpain.     

The stimulatory effect of beta-endorphin on the plasma prolactin levels was diminished in the rats treated with 6-hydroxydopamine

Intraventicular injection of beta-endorphin (beta LPH_61^?91) in urethane anesthetized male rats led to a dose dependent increase of plasma prolactin levels. Intravenous injection of apomorphine completely abolished the stimulatory effect of beta-endorphin. Animals treated with 6-hydroxydopamine (6-OHDA) and 6-OHDA plus desmethylimipramine showed inhibition of beta-endorphin induced prolactin release. These results suggest that beta-endorphin presynaptically inhibits the activity of dopaminergic neurones, leading to the stimulation of plasma prolactin levels.     

Structural characterization and dynamics of globotetraosylceramide in vascular endothelial cells under TNF-α stimulation

In several vascular inflammatory reactions (i.e. immunity and thrombosis) inflammatory mediators lead to the activation of vascular endothelial cells (EC). To date, a number of functional molecules induced on the surface of activated-EC have been identified. We report here that Globotetraosylceramide (Gb4), a glycosphingolipid expressed in EC, is a novel inducible molecule on EC activated by TNF-α. The cell surface expression of Gb4 is increased in a time-dependent manner under TNF-α stimulation, which shows distinct expression kinetics of major proteins induced by TNF-α on EC. MALDI-TOF-MS analysis revealed that the enhanced Gb4 predominantly contains C24:0 fatty acid in the ceramide moiety. Isolated caveolae/lipid raft-enriched detergent insoluble membrane domains in activated-EC predominantly contain this molecular species of Gb4. Gb4 containing C16:0 fatty acid in the ceramide moiety, which is known to constitute the major species of Gb4 in plasma, is also found as a major molecular species in EC. These observations indicate that Gb4, especially with very long fatty acid, is enhanced in EC during its inflammatory reaction, and suggest the potential utility of Gb4 as a biomarker for monitoring inflammation status of EC involving its related diseases.     

A simple and efficient synthesis of puromycin, 2,2′-anhydro-pyrimidine nucleosides,cytidines and 2′,3′-anhydroadenosine from 3′,5′-O-sulfinyl Xylo-nucleosides

Synthesis of antibiotics, puromycin and 3 ′-amino-3 ′-deoxy-N ⁶,N ⁶-dimethyladenosine 11 was achieved by utilizing the cyclic sulfite 6a of the xylo-3 ′,5 ′-dihydroxy group as a new protective group. The key synthetic step is the deprotection of the sulfite moiety through the intramolecular cyclization of 2-α-carbamate 7. In a similar manner, 2,2 ′-anhydro-pyrimidine nucleosides 15, ribo-cytidines 17 and 2 ′,3 ′-anhydroadenosine 14 were prepared in high yields from the corresponding sulfites 4, 5, and 6b, respectively.     

Molecular Structure of Xanthocidin

In order to establish industrial production of 5′-inosinic acid (5′-IMP), a permeability mutant, KY13171, of Brevibacterium ammoniagenes, which accumulated 7 to 8 grams of 5′-IMP per liter and 4 to 6 grams of hypoxanthine (Hx) per liter (calculated as 5′-IMP), was improved by a genetical procedure. Further improved mutants were selected stepwise through repeating mutational work. The finally selected mutant. KY13369, accumulated 20 to 27 grams of 5′-IMP per liter, but not Hx.

Increased productivity of 5′-IMP and decreased productivity of Hx were not caused by the changes in 5′-IMP degrading activity, because these activities were not significantly different among the mutants. These results appear to indicate that the increased accumulation of 5′-IMP may be caused by the improvement in membrane permeability for 5′-IMP. However, the changes in phospholipid and fatty acid compositions were not enough to explain the increased permeability.     

Identification and estimation algorithm for stochastic neural system. II

The algorithm for identifying the stochastic neural system and estimating the system process which reflects the dynamics of the neural network are presented in this papar. The analogous algorithm has been proposed in our preceding paper (Nakao et al., 1984), which was based on the randomly missed observations of a system process only. Since the previous algorithm mentioned above was subject to an unfavorable effect of consecutively missed observations, to reduce such an effect the algorithm proposed here is designed additionally to observe an intensity process in a neural spike train as the information for the estimation.The algorithm is constructed with the extended Kalman filters because it is naturally expected that a nonlinear and time variant structure is necessary for the filters to realize the observation of an intensity process by means of mapping from a system process to an intensity process. The performance of the algorithm is examined by applying it to some artificial neural systems and also to cat's visual nervous systems. The results in these applications are thought to prove the effectiveness of the algorithm proposed here and its superiority to the algorithm proposed previously.