Pan Dong Ryu

Pre- and post-treatments with escitalopram protect against experimental ischemic neuronal damage via regulation of BDNF expression and oxidative stress

Selective serotonin re-uptake inhibitors (SSRI) have been widely used in treatment of major depression because of their efficacy, safety, and tolerability. Escitalopram, an SSRI, is known to decrease oxidative stress in chronic stress animal models. In the present study, we examined the neuroprotective effects of pre- and post-treatments with 20 mg/kg and 30 mg/kg escitalopram in the gerbil hippocampal CA1 region (CA1) after transient cerebral ischemia. Pre-treatment with escitalopram protected against ischemia-induced neuronal death in the CA1 after ischemia/reperfusion (I/R). Post-treatment with 30 mg/kg, not 20 mg/kg, escitalopram had a neuroprotective effect against ischemic damage. In addition, 20 mg/kg pre- and 30 mg/kg post-treatments with escitalopram increased brain-derived neurotrophic factor (BDNF) protein levels in the ischemic CA1 compared to vehicle-treated ischemia animals. In addition, 20 mg/kg pre- and 30 mg/kg post-treatments with escitalopram reduced microglia activation and decreased 4-hydroxy-2-nonenal and Cu,Zn-superoxide dismutase immunoreactivity and their levels in the ischemic CA1 compared to vehicle-treated ischemia animals after transient cerebral ischemia. In conclusion, these results indicated that pre- and post-treatments with escitalopram can protect against ischemia-induced neuronal death in the CA1 induced by transient cerebral ischemic damage by increase of BDNF as well as decrease of microglia activation and oxidative stress.

Human umbilical cord blood-derived mesenchymal stem cells protect against neuronal cell death and ameliorate motor deficits in Niemann Pick type C1 mice.

Niemann Pick disease type C1 (NPC) is an autosomal recessive disease characterized by progressive neurological deterioration leading to premature death. In this study, we hypothesized that human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) have the multifunctional abilities to ameliorate NPC symptoms in the brain. To test this hypothesis, hUCB-MSCs were transplanted into the hippocampus of NPC mice in the early asymptomatic stage. This transplantation resulted in the recovery of motor function in the Rota Rod test and impaired cholesterol homeostasis leading to increased levels of cholesterol efflux-related genes such as LXRα, ABCA1, and ABCG5 while decreased levels of 3-hydroxy-3-methylglutaryl coenzyme A reductase were observed in NPC mice. In the cerebrum, hUCB-MSCs enhanced neuronal cell survival and proliferation, where they directly differentiated into electrically active MAP2-positive neurons as demonstrated by whole-cell patch clamping. In addition, we observed that hUCB-MSCs reduced Purkinje neuronal loss by suppression of inflammatory and apoptotic signaling in the cerebellum as shown by immunohistochemistry. We further investigated how hUCB-MSCs enhance cellular survival and inhibit apoptosis in NPC mice. Neuronal cell survival was associated with increased PI3K/AKT and JAK2/STAT3 signaling; moreover, hUCB-MSCs modulated the levels of GABA/glutamate transporters such as GAT1, EAAT2, EAAT3, and GAD6 in NPC mice as assessed by Western blot analysis. Taken together, our findings suggest that hUCB-MSCs might play multifunctional roles in neuronal cell survival and ameliorating motor deficits of NPC mice.

Tonic Extrasynaptic GABAA Receptor Currents Control Gonadotropin-Releasing Hormone Neuron Excitability in the Mouse

It is well established that the GABA(A) receptor plays an important role in regulating the electrical excitability of GnRH neurons. Two different modes of GABA(A) receptor signaling exist: one mediated by synaptic receptors generating fast (phasic) postsynaptic currents and the other mediated by extrasynaptic receptors generating a persistent (tonic) current. Using GABA(A) receptor antagonists picrotoxin, bicuculline methiodide, and gabazine, which differentiate between phasic and tonic signaling, we found that ∼50% of GnRH neurons exhibit an approximately 15-pA tonic GABA(A) receptor current in the acute brain slice preparation. The blockade of either neuronal (NO711) or glial (SNAP-5114) GABA transporter activity within the brain slice revealed the presence of tonic GABA signaling in ∼90% of GnRH neurons. The GABA(A) receptor δ subunit is only found in extrasynaptic GABA(A) receptors. Using single-cell RT-PCR, GABA(A) receptor δ subunit mRNA was identified in GnRH neurons and the δ subunit-specific agonist 4,5,6,7-tetrahydroisoxazolo [5,4-c] pyridin-3-ol was found to activate inward currents in GnRH neurons. Perforated-patch clamp studies showed that 4,5,6,7-tetrahydroisoxazolo [5,4-c] pyridin-3-ol exerted the same depolarizing or hyperpolarizing effects as GABA on juvenile and adult GnRH neurons and that tonic GABA(A) receptor signaling regulates resting membrane potential. Together, these studies reveal the presence of a tonic GABA(A) receptor current in GnRH neurons that controls their excitability. The level of tonic current is dependent, in part, on neuronal and glial GABA transporter activity and mediated by extrasynaptic δ subunit-containing GABA(A) receptors.

Reduction in synaptic GABA release contributes to target-selective elevation of PVN neuronal activity in rats with myocardial infarction

Neuronal activity in the paraventricular nucleus (PVN) is known to be elevated in rats with heart failure. However, the type of neurons involved and the underlying synaptic mechanisms remain unknown. Here we examined spontaneous firing activity and synaptic currents in presympathetic PVN neurons in rats with myocardial infarction (MI), using slice patch clamp combined with the retrograde labeling technique. In PVN neurons projecting to the rostral ventrolateral medulla (PVN-RVLM), MI induced a significant increase in basal firing rate (1.79 to 3.02 Hz, P < 0.05) and a reduction in the frequency of spontaneous (P < 0.05) and miniature (P < 0.01) inhibitory postsynaptic currents (IPSCs). In addition, MI induced an increase in the paired-pulse ratio of evoked IPSCs (P < 0.05). Bicuculline, a GABA(A) receptor antagonist, increased the firing rate of PVN-RVLM neurons in sham-operated (1.21 to 2.74 Hz, P < 0.05) but not MI (P > 0.05) rats. In contrast, in PVN neurons projecting to the intermediolateral horn of the spinal cord (PVN-IML), MI did not induce any significant changes in the basal firing rate and the properties of spontaneous and miniature IPSCs. The properties of spontaneous excitatory postsynaptic currents (EPSCs) were not altered in either neuron group. In conclusion, our results indicate that MI induces an elevation of firing activity in PVN-RVLM but not in PVN-IML neurons and that the elevated firing rate is largely due to a decrease in GABA release. These results provide evidence for a novel target-selective synaptic plasticity in the PVN that is associated with the sympathetic hyperactivity commonly seen in heart failure.

Gold nanoparticles promote osteogenic differentiation in human adipose-derived mesenchymal stem cells through the Wnt/β-catenin signaling pathway.

Gold nanoparticles (AuNPs) are attractive materials for use in biomedicine due to their physical properties. Increasing evidence suggests that several nanoparticles induce the differentiation of human mesenchymal stem cells into osteoblasts and adipocytes. In this study, we hypothesized that chitosan-conjugated AuNPs promote the osteogenic differentiation of human adipose-derived mesenchymal stem cells. For the evaluation of osteogenic differentiation, alizarin red staining, an alamarBlue® assay, and a quantitative real-time polymerase chain reaction analysis were performed. In order to examine specific signaling pathways, immunofluorescence and a western blotting assay were performed. Our results demonstrate that chitosan-conjugated AuNPs increase the deposition of calcium content and the expression of marker genes related to osteogenic differentiation in human adipose-derived mesenchymal stem cells at nontoxic concentrations. These results indicate that chitosan-conjugated AuNPs promote osteogenesis through the Wnt/β-catenin signaling pathway. Therefore, chitosan-conjugated AuNPs can be used as a reagent for promoting bone formation.

Activation of presynaptic group I metabotropic glutamate receptors enhances glutamate release in the rat spinal cord substantia gelatinosa.

The activation of group I metabotropic glutamate receptors (mGluRs) produces a long-term potentiation of sensory transmission in the substantia gelatinosa (SG) region of the spinal cord (Prog. Brain Res. 129 (2000) 115). The mechanism(s) responsible for the induction of this potentiation is not known. Using rat spinal cord slice preparation and patch-clamp recordings, here we show, that the activation of the group I mGluRs by (S)-3,5-dihydroxyphenylglycine (DHPG, 1 microM), the mGluR1/5 agonist, increased the frequency of both activity-dependent spontaneous EPSCs, and activity-independent miniature EPSCs (mEPSCs). However, DHPG did not affect amplitude of mEPSCs. The effects of DHPG were not seen in the presence of the preferential mGluR1 antagonist CPCCOEt (10 microM). On the other hand, 2-methyl-6-(phenylethynyl)-pyridine (10 microM), a selective mGluR5 antagonist, blocked the DHPG facilitation present during the wash-out of the drug. This novel facilitating effect of the group I mGluR activation on glutamate release is the first report of a direct facilitatory action of both mGluR1 and mGluR5 subtypes on sensory transmission in the spinal cord SG region. These results indicate the potential contribution of synaptic activation of these facilitatory autoreceptors in plasticity of primary afferent neurotransmission.

Cation selective channels formed by a C-terminal fragment of β-amyloid precursor protein

The C-terminal 105 amino acid fragment of beta-amyloid precursor protein (CT105) is highly neurotoxic. To obtain insights into its cytotoxic effect, we examined the ionophoric effects of CT105 (10-1000 nM) on artificial lipid membranes. Macroscopic membrane conductance increased with CT105 concentration and its ionophoric effect was comparable to that of amyloid beta protein. The mean unitary conductance of CT105-induced channels was 120 pS and open-state probability was close to 1 at voltages from -80 to +80 mV. CT105induced channels were selective to cations (PK/ P(Cl) = 10.2), being most selective to Ca2+. These findings suggest that CT105 can cause direct neurotoxic effects by forming Ca2+ permeable cation channels on neuronal membranes.

Imbalanced K+ and Ca2+ subthreshold interactions contribute to increased hypothalamic presympathetic neuronal excitability in hypertensive rats

Despite the importance of brain‐mediated sympathetic activation in the morbidity and mortality of patients with high blood pressure, the precise cellular mechanisms involved remain largely unknown. We show that an imbalanced interaction between two opposing currents mediated by potassium (IA) and calcium (IT) channels occurs in sympathetic‐related hypothalamic neurons in hypertensive rats. We show that this imbalance contributes to enhanced membrane excitability and firing activity in this neuronal population. Knowledge of how these opposing ion channels interact in normal and disease states increases our understanding of underlying brain mechanisms contributing to the high blood pressure condition.

Voltage-gated K+ channels in adipogenic differentiation of bone marrow-derived human mesenchymal stem cells

Aim:To determine the presence of voltage-gated K+ (Kv) channels in bone marrow-derived human mesenchymal stem cells (hMSCs) and their impact on differentiation of hMSCs into adipocytes.Methods:For adipogenic differentiation, hMSCs were cultured in adipogenic medium for 22 d. The degrees of adipogenic differentiation were examined using Western blot, Oil Red O staining and Alamar assay. The expression levels of Kv channel subunits Kv1.1, Kv1.2, Kv1.3, Kv1.4, Kv2.1, Kv3.1, Kv3.3, Kv4.2, Kv4.3, and Kv9.3 in the cells were detected using RT-PCR and Western blot analysis.Results:The expression levels of Kv2.1 and Kv3.3 subunits were markedly increased on d 16 and 22. In contrast, the expression levels of other Kv channel subunits, including Kv1.1, Kv1.2, Kv1.3, Kv1.4, Kv4.2, Kv4.3, and Kv9.3, were decreased as undifferentiated hMSCs differentiated into adipocytes. Addition of the Kv channel blocker tetraethylammonium (TEA, 10 mmol/L) into the adipogenic medium for 6 or 12 d caused a significant decrease, although not complete, in lipid droplet formation and adipocyte fatty acid-binding protein 2 (aP2) expressions. Addition of the selective Kv2.1 channel blocker guangxitoxin (GxTX-1, 40 nmol/L) into the adipogenic medium for 21 d also suppressed adipogenic differentiation of the cells.Conclusion:The results demonstrate that subsets of Kv channels including Kv2.1 and Kv3.3 may play an important role in the differentiation of hMSCs into adipocytes.

Possible presence of the ATP-sensitive K+ channel in isolated spinal dorsal horn neurons of the rat

The ATP-sensitive K+ channel (KATP channel) is a K+ channel inhibited by cytoplasmic ATP. It was originally found in cardiac cells and recently in neuronal cells. Here, we present evidence indicating that the KATP channel also exists in spinal dorsal horn neurons: membrane currents were recorded by whole-cell voltage-clamp in spinal dorsal horn neurons isolated from young rats. The outward current was augmented by KATP channel activators nicorandil and minoxidil and reduced by the blocker glibenclamide. This glibenclamide-induced change in the current was augmented when the intracellular ATP was lowered and the reversal potential was shifted according to the external K+ concentration.