TGF-beta1 suppresses T cell infiltration and VP2 puff B mutation enhances apoptosis in acute polioencephalitis induced by Theiler's virus

GDVII and DA strains of Theiler's murine encephalomyelitis virus (TMEV) differ in VP2 puff B. One week after GDVII virus infection, SJL/J mice had large numbers of TUNEL+ apoptotic cells with a relative lack of T cell infiltration in the brain. DA viruses with mutation in puff B induced higher levels of apoptosis than wild-type DA virus, but levels of inflammation in brains were similar between DA and DA virus mutants. The difference in inflammation among TMEVs could be due to TGF-beta1 expression that was seen only in GDVII virus infection and negatively correlated with CD3+ T cell infiltration.     

Enriched environments influence depression-related behavior in adult mice and the survival of newborn cells in their hippocampi

Major depression is a highly prevalent mental disorder and environmental factors have been strongly implicated in its pathophysiology. Clinical studies have demonstrated that stress or depression can lead to atrophy and cell loss in the hippocampus. Studies of animal models of depression have suggested that reduced neurogenesis in the adult hippocampus might contribute to such structural changes and to the behavior of these animals. On the other hand, increased hippocampal neurogenesis can be induced by the administration of antidepressants or electroconvulsive seizure, suggesting that increased neurogenesis might be related to the treatment of depression. Thus, an enriched environment (EE), which also enhances neurogenesis, is expected to have therapeutic effects on depression-related behaviors. To investigate the effects of an EE during adulthood on these behaviors, we subjected adult mice housed in an EE for five weeks to behavioral tests. In an open field test, EE mice exhibited a decrease in the distance traveled and an increase in the amount of time spent in the center. The startle response was smaller in EE mice than in control mice. EE mice also showed reduced immobility time in a forced swim test. The immobility time in EE mice was approximately half that observed in mice treated with a tricyclic antidepressant, imipramine. In our experimental condition, increased survival of newborn cells was observed in EE mice by 5-bromo-2'-deoxyuridine (BrdU)-labeled immunohistochemistry. Double-staining of BrdU and a mature neuron marker, NeuN, revealed that the majority of surviving cells were neurons. Our results suggest that EE, which enhanced the survival of newborn neurons, shows beneficial effects on behavioral despair and habituation to a novel environment.     

Topological and chronological features of the impairment of glucose metabolism induced by 1-methyl-4-phenylpyridinium ion (MPP<Superscript>+</Superscript>) in rat brain slices

Summary 1-Methyl-4-phenylpyridinium (MPP⁺) was added directly to fresh rat brain slices and the dynamic changes in the cerebral glucose metabolic rate (CMRglc) were serially and two-dimensionally measured with [¹⁸F]2-fluoro-2-deoxy-D-glucose as a tracer. MPP⁺ dose-dependently increased CMRglc, reflecting enhanced glycolysis compensating for the decrease in aerobic metabolism. While the CMRglc enhancement induced by MPP⁺ (<10 μM) was restricted to the striatum, MPP⁺ (≥10 μM) induced a significant CMRglc enhancement in all brain regions. MPP⁺ at high concentration (1 mM) eventually initiated rapid metabolic collapse, with failure to sustain anaerobic glycolysis.     

PACAP/PAC1 autocrine system promotes proliferation and astrogenesis in neural progenitor cells

The Pituitary adenylate cyclase-activating peptide (PACAP) ligand/type 1 receptor (PAC1) system regulates neurogenesis and gliogenesis. It has been well established that the PACAP/PAC1 system induces differentiation of neural progenitor cells (NPCs) through the Gs-mediated cAMP-dependent signaling pathway. However, it is unknown whether this ligand/receptor system has a function in proliferation of NPCs. In this study, we identified that PACAP and PAC1 were highly expressed and co-localized in NPCs of mouse cortex at embryonic day 14.5 (E14.5) and found that the PACAP/PAC1 system potentiated growth factor-induced proliferation of mouse cortical NPCs at E14.5 via Gq-, but not Gs-, mediated PLC/IP3-dependent signaling pathway in an autocrine manner. Moreover, PAC1 activation induced elongation of cellular processes and a stellate morphology in astrocytes that had the bromodeoxyuridine (BrdU)-incorporating ability of NPCs. Consistent with this notion, we determined that the most BrdU positive NPCs differentiated to astrocytes through PAC1 signaling. These results suggest that the PACAP/PAC1 system may play a dual role in neural/glial progenitor cells not only differentiation but also proliferation in the cortical astrocyte lineage via Ca2+-dependent signaling pathways through PAC1.     

Drug Development Targeting the Glycogen Synthase Kinase-3β (GSK-3β)-Mediated Signal Transduction Pathway: The Role of GSK-3β in the Maintenance of Steady-State Levels of Insulin Receptor Signaling Molecules and Nav1.7 Sodium Channel in Adrenal Chromaffin Cells

Glycogen synthase kinase-3 (GSK-3) is constitutively active in nonstimulated cells, where the majority of its substrates undergo inactivation/proteolysis by phosphorylation. Extracellular stimuli (e.g., insulin) catalyze inhibitory Ser⁹-phosphorylation of GSK-3β, turning on signaling and causing other biological consequences otherwise constitutively suppressed by GSK-3β. Regulated and dysregulated activities of GSK-3β are pivotal to health, disease, and therapeutics (e.g., insulin resistance, neurodegeneration, tumorigenesis, inflammation); however, the underlying mechanisms of multifunctional GSK-3β remain elusive. In cultured bovine adrenal chromaffin cells, 1) constitutive and negatively-regulated activities of GSK-3β up- and down-regulated insulin receptor, insulin receptor substrate-1 (IRS-1), IRS-2, and Akt levels via controlling proteasomal degradation and protein synthesis; 2) nicotinic receptor/protein kinase C-α (PKC-α) / extracellular signal-regulated kinase (ERK) pathway up-regulated IRS-1 and IRS-2 levels, enhancing insulin-induced the phosphoiNOSitide 3-kinase (PI3K) / Akt / GSK-3β pathway; 3) inhibition of calcineurin by cyclosporin A or FK506 down-regulated IRS-2 level, attenuating insulin-like growth factor-I (IGF-I)-induced ERK and GSK-3β pathways; and 4) insulin, IGF-I or therapeutics (e.g., lithium) up-regulated the voltage-dependent Na_v1.7 sodium channel.     

Wortmannin Inhibits the Increase in Myofilament Ca2+ Sensitivity Induced by Cross-Talk of Endothelin-1 With Norepinephrine in Canine Ventricular Myocardium

Endothelin-1 (ET-1) modulates cardiac contractility by cross-talk with norepinephrine (NE) in canine ventricular myocardium. The present experiments were performed to investigate the influence of wortmannin that has inhibitory action on phosphatidylinositol 3-kinase (PI3-K) (IC₅₀ = 3 nM) and myosin light chain kinase (MLCK) (IC₅₀ = 200 nM) on Ca²⁺ signaling and the inotropic effects of ET-1 induced by cross-talk with NE. Experiments were carried out in isolated canine ventricular trabeculae and indo-1/AM–loaded single ventricular cardiomyocytes. ET-1 alone elicited a transient small negative inotropic effect (NIE). In the presence of NE at low (1 – 10 nM) and high (100 nM) concentrations, ET-1 induced a long-lasting positive inotropic effect (PIE) or a marked sustained NIE, respectively. Wortmannin up to 300 nM did not affect the contractility; and at 1 μM and higher, it decreased the basal contraction without suppressing Ca²⁺ transients. Wortmannin (1 μM) inhibited the long-lasting PIE of ET-1 without affecting the ET-1–induced increase in Ca²⁺ transients. Wortmannin at the same concentration did not affect the ET-1–induced transient and sustained NIE and the PIE mediated by β-adrenoceptor stimulation. These results imply that wortmannin exerts selective inhibitory action on the increase in myofilament Ca²⁺ sensitivity induced by cross-talk of ET-1 with NE probably through an inhibition of MLCK in canine ventricular myocardium.     

Development of novel mucoadhesive pellets of metformin hydrochloride

Mucoadhesive polymer-coated pellets containing metformin hydrochloride were prepared by the powder-layering technique using a centrifugal fluidizing (CF)-granulator. Four high-viscosity polymers were applied to make the pellets: 1) hydroxymethylcellulose (HPMC), 2) sodium alginate (Na-Alg), 3) HPMC/Carbopol, and 4) sodium carboxylmethylcellulose (Na-CMC). The physical crushing test, mucoadhesive test, zeta-potential test, in vitro release study and observation of gastroretention state of the dosage form were performed to investigate the pellets. The strong adhesive interaction between the Na-CMC-coated pellets and the mucin disc was obtained by mucoadhesive test. Na-Alg was most effective among the polymers used in changing the value of zeta potential of the mucin solution by the interaction between a polymer and a mucin particle. Results from drug dissolution study showed that over 95% of the drug from all the four pellets was released before 2 h, while Na-CMC- and Na-Alg-coated pellets showed a moderate sustained-release in SGF (simulated gastric fluid) and SIF (simulated intestine fluid), respectively. In conclusion, Na-CMC and Na-Alg seem to be promising candidates for mucoadhesive formulation and further studies to improve the sustained-release property are underway for achieving the ultimate goal of once-a-day formulation of metformin hydrochloride.