Naohisa Arakawa

Na+–Ca2+ exchanger isoforms in rat neuronal preparations: different changes in their expression during postnatal development

We examined the relative amounts of Na(+)-Ca(2+) exchanger (NCX) isoform mRNAs in cultured neurons, astrocytes and developmental rat brain. NCX1 transcript was predominant in neurons and astrocytes, but NCX2 transcript was about four-fold higher than NCX1 or NCX3 transcript in adult rat cortex. NCX2 transcript in the cortex increased markedly during postnatal development, whereas NCX1 and NCX3 transcripts decreased. Na(+)-dependent 45Ca(2+) uptake in the cortical homogenate increased significantly during postnatal development.

Differential expression of mRNAs for PACAP and its receptors during neural differentiation of embryonic stem cells.

The expressions of mRNAs for pituitary adenylate cyclase-activating polypeptide (PACAP), vasoactive intestinal peptide (VIP), and their receptors (PAC1, VPAC1 and VPAC2) were examined in the five steps of the in vitro neuronal culture model of embryonic stem (ES) cell differentiation. mRNAs for PACAP, VIP, PAC1 receptor, and VPAC2 receptor were moderately expressed in neural stem cell-enriched cultures, while VPAC1 receptor mRNA was most prominently expressed in embryoid bodies (EBs). The expression of PAC1 receptor mRNA was further upregulated after terminal differentiation into neurons. In contrast, the expressions of PAC1 receptor and PACAP mRNAs were markedly decreased after glial differentiation. These results suggest that this in vitro neuronal culture system will be a useful model for future studies on the functional role of the PACAPergic system during different stages of neuronal development.

Inhibition of Self‐Renewal and Induction of Neural Differentiation by PACAP in Neural Progenitor Cells

Abstract:  Several lines of evidence have suggested roles for pituitary adenylate cyclase‐activating polypeptide (PACAP) in the developing nervous system. Previously, we showed that mRNA for PACAP, vasoactive intestinal peptide (VIP), and their three receptor subtypes, is differentially expressed in embryonic stem (ES) cells, ES cell‐derived, neural stem cell‐enriched cultures, and differentiated neurons, by using the five steps of the in vitro neuronal culture model of ES cell differentiation . Here, we examined the effects of PACAP on self‐renewal and cell lineage determination of neural progenitor/stem cells. PACAP inhibited the basic fibroblast growth factor‐induced proliferation (self‐renewal), as assessed by neurosphere formation. PACAP increased microtubule‐associated protein 2‐positive neurons without affecting the number of cells positive for the neural stem cell marker nestin, astrocyte marker glial fibrillary acidic protein, and oligodendrocyte marker CNPase. These results suggest that PACAP inhibits self‐renewal but, instead, induces early neuronal differentiation of neural progenitor cells.

Role of Angiotensin II Type 1 Receptor on Retinal Vascular Leakage in a Rat Oxygen-Induced Retinopathy Model

Aim: To investigate the role of angiotensin type 1 (AT1) and type 2 (AT2) receptors in hypoxia-induced retinal vascular hyperpermeability. Methods: Brown-Norway rat pups were exposed to hyperoxic conditions from postnatal day 7 (P7) to P12, and to subsequent normal air for 5 days [oxygen-induced retinopathy (OIR) model]. Olmesartan medoxomil (AT1 receptor antagonist; administered orally), PD123319 (AT2 receptor antagonist; administered subcutaneously) or a vehicle was administered once daily during the last 5 days. At P16, the retinal permeability was determined by measuring the leaked fluorescein-conjugated dextran concentration in the retina. The vascular endothelial growth factor (VEGF) and hypoxia-inducible factor 1 (HIF-1) α proteins in the retina were assessed by an ELISA and western blotting, respectively. Results: Olmesartan medoxomil partially, but significantly, inhibited the retinal vascular hyperpermeability induced by hypoxia. In contrast, PD123319 did not show a significant effect. The VEGF and HIF-1α protein levels were significantly elevated in the OIR retina; however, there was no significant effect of olmesartan medoxomil on the expression of either protein. Conclusions: These results suggest that the AT1 receptor is, at least partly, responsible for hyperpermeability in the OIR rat retina via a mechanism independent of HIF-1 and VEGF expression.

The Na+-Ca2+ exchange inhibitor KB-R7943 inhibits high K+-induced increases in intracellular Ca2+ concentration and [3H]noradrenaline release in the human neuroblastoma SH-SY5Y.

The effects of the Na+-Ca2+ exchange inhibitor 2-[2-[4-(4-nitrobenzyloxy)phenyl]ethyl]isothiourea methanesulfonate (KB-R7943) on depolarization-induced Ca2+ signal and [3H]noradrenaline release were examined in SH-SY5Y cells. KB-R7943 at 10 μM significantly inhibited high K+-induced increase in intracellular Ca2+ concentration. KB-R7943 also inhibited high K+-evoked release of [3H]noradrenaline from the cells. These findings suggest that the Na+-Ca2+ exchanger in the reverse mode is involved at least partly in depolarization-induced transmitter release.

Olmesartan medoxomil ameliorates sciatic nerve regeneration in diabetic rats.

To evaluate the effect of angiotensin II type1 receptor blocker on nerve regeneration delay in diabetic rats, nerve regeneration was monitored by a pinch test on the crushed sciatic nerves of streptozotocin-induced diabetic rats. Nerve regeneration was significantly delayed in diabetic rats and was partly ameliorated by treatment with olmesartan medoxomil (3 mg/kg/day, orally). In the ipsilateral dorsal root ganglia, the mRNA level of insulin-like growth factor-1 and ciliary neurotrophic factor (CNTF) was downregulated, whereas the mRNA level of neurotrophin-3 and CNTF receptor was upregulated. Olmesartan medoxomil significantly enhanced the CNTF expression. These results showed that angiotensin II type1 receptor blocker treatment is effective on nerve regeneration delay in diabetic animals and may provide an effective therapy for clinical diabetic neuropathy.

Alkylsulfanyl analogs as potent α 2 δ ligands

We identified novel (3R, 5S)-3-aminomethyl-5-methanesulfanyl hexanoic acid (5a: DS75091588) and (3R, 5S)-3-aminomethyl-5-ethanesulfanyl hexanoic acid (6a: DS18430756) as sulfur-containing γ-amino acid derivatives that were useful for the treatment of neuropathic pain. These two compounds exhibited a potent analgesic effect in animal models of both type I diabetes and type II diabetes, and good pharmacokinetics.

Contents Vol. 41, 2009

Anatomy, Pathology and Cell Biology A. Prescott, Dundee Biochemistry, Molecular Biology and Molecular Genetics J. Graw, Neuherberg Clinical and Epidemiological Research M. Kojima, Kahoku Clinical Retina P. Wiedemann, Leipzig Cornea and Ocular Surface C. Marfurt, Gary, Ind. Glaucoma C. Erb, Berlin Immunology and Microbiology U. Pleyer, Berlin Lens and Cataract S. Varma, Baltimore, Md. Miscellaneous U. Pleyer, Berlin Neuro-Ophthalmology and Vision Sciences P. Aydin, Ankara Ocular Oncology M. Jager, Leiden Physiology, Pharmacology and Toxicology A. Wegener, Bonn Retina and Retinal Cell Biology M. Boulton, Gainesville, Fla. P. Wiedemann, Leipzig Editorial Board