Sonic hedgehog signalling mediates astrocyte crosstalk with neurons to confer neuroprotection.

Sonic hedgehog (SHH) is a glycoprotein associated with development that is also expressed in the adult CNS and released after brain injury. Since the SHH receptors patched homolog-1 and Smoothened are highly expressed on astrocytes, we hypothesized that SHH regulates astrocyte function. Primary mouse cortical astrocytes derived from embryonic Swiss mouse cortices, were treated with two chemically distinct agonists of the SHH pathway, which caused astrocytes to elongate and proliferate. These changes are accompanied by decreases in the major astrocyte glutamate transporter-1 and the astrocyte intermediate filament protein glial fibrillary acidic protein. Multisite electrophysiological recordings revealed that the SHH agonist, smoothened agonist suppressed neuronal firing in astrocyte-neuron co-cultures and this was abolished by the astrocyte metabolic inhibitor ethylfluoroacetate, revealing that SHH stimulation of metabolically active astrocytes influences neuronal firing. Using three-dimensional co-culture, MAP2 western blotting and immunohistochemistry, we show that SHH-stimulated astrocytes protect neurons from kainate-induced cell death. Altogether the results show that SHH regulation of astrocyte function represents an endogenous neuroprotective mechanism.

Cannabidiol displays antiepileptiform and antiseizure properties in vitro and in vivo.

Plant-derived cannabinoids (phytocannabinoids) are compounds with emerging therapeutic potential. Early studies suggested that cannabidiol (CBD) has anticonvulsant properties in animal models and reduced seizure frequency in limited human trials. Here, we examine the antiepileptiform and antiseizure potential of CBD using in vitro electrophysiology and an in vivo animal seizure model, respectively. CBD (0.01-100 muM) effects were assessed in vitro using the Mg(2+)-free and 4-aminopyridine (4-AP) models of epileptiform activity in hippocampal brain slices via multielectrode array recordings. In the Mg(2+)-free model, CBD decreased epileptiform local field potential (LFP) burst amplitude [in CA1 and dentate gyrus (DG) regions] and burst duration (in all regions) and increased burst frequency (in all regions). In the 4-AP model, CBD decreased LFP burst amplitude (in CA1 only at 100 muM CBD), burst duration (in CA3 and DG), and burst frequency (in all regions). CBD (1, 10, and 100 mg/kg) effects were also examined in vivo using the pentylenetetrazole model of generalized seizures. CBD (100 mg/kg) exerted clear anticonvulsant effects with significant decreases in incidence of severe seizures and mortality compared with vehicle-treated animals. Finally, CBD acted with only low affinity at cannabinoid CB(1) receptors and displayed no agonist activity in [(35)S]guanosine 5'-O-(3-thio)triphosphate assays in cortical membranes. These findings suggest that CBD acts, potentially in a CB(1) receptor-independent manner, to inhibit epileptiform activity in vitro and seizure severity in vivo. Thus, we demonstrate the potential of CBD as a novel antiepileptic drug in the unmet clinical need associated with generalized seizures.

Δ9-Tetrahydrocannabivarin suppresses in vitro epileptiform and in vivo seizure activity in adult rats.

PURPOSE: We assessed the anticonvulsant potential of the phytocannabinoid Δ9-tetrahydrocannabivarin (Δ9-THCV) by investigating its effects in an in vitro piriform cortex (PC) brain slice model of epileptiform activity, on cannabinoid CB1 receptor radioligand-binding assays and in a generalized seizure model in rats. METHODS: Δ9-THCV was applied before (10 µm Δ9-THCV) or during (10-50 µm Δ9-THCV) epileptiform activity induced by Mg²(+) -free extracellular media in adult rat PC slices and measured using multielectrode array (MEA) extracellular electrophysiologic techniques. The actions of Δ9-THCV on CB1 receptors were examined using [³H]SR141716A competition binding and [³5S]GTPγS assays in rat cortical membranes. Effects of Δ9-HCV (0.025-2.5 mg/kg) on pentylenetetrazole (PTZ)-induced seizures in adult rats were also assessed. RESULTS: After induction of stable spontaneous epileptiform activity, acute Δ9 -THCV application (≥ 20 µm) significantly reduced burst complex incidence and the amplitude and frequency of paroxysmal depolarizing shifts (PDSs). Furthermore, slices pretreated with 10 µm Δ9-THCV prior to induction of epileptiform activity exhibited significantly reduced burst complex incidence and PDS peak amplitude. In radioligand-binding experiments, Δ9-THCV acted as a CB1 receptor ligand, displacing 0.5 nm [³H]SR141716A with a Ki∼290 nm, but exerted no agonist stimulation of [³5S]GTPγS binding. In PTZ-induced seizures in vivo, 0.25 mg/kg Δ9-THCV significantly reduced seizure incidence. DISCUSSION: These data demonstrate that Δ9-THCV exerts antiepileptiform and anticonvulsant properties, actions that are consistent with a CB1 receptor-mediated mechanism and suggest possible therapeutic application in the treatment of pathophysiologic hyperexcitability states.

Human neural stem cell-derived cultures in three-dimensional substrates form spontaneously functional neuronal networks.

Differentiated human neural stem cells were cultured in an inert three-dimensional (3D) scaffold and, unlike two-dimensional (2D) but otherwise comparable monolayer cultures, formed spontaneously active, functional neuronal networks that responded reproducibly and predictably to conventional pharmacological treatments to reveal functional, glutamatergic synapses. Immunocytochemical and electron microscopy analysis revealed a neuronal and glial population, where markers of neuronal maturity were observed in the former. Oligonucleotide microarray analysis revealed substantial differences in gene expression conferred by culturing in a 3D vs a 2D environment. Notable and numerous differences were seen in genes coding for neuronal function, the extracellular matrix and cytoskeleton. In addition to producing functional networks, differentiated human neural stem cells grown in inert scaffolds offer several significant advantages over conventional 2D monolayers. These advantages include cost savings and improved physiological relevance, which make them better suited for use in the pharmacological and toxicological assays required for development of stem cell-based treatments and the reduction of animal use in medical research. Copyright © 2015 John Wiley & Sons, Ltd.

Neuronal-glial populations form functional networks in a biocompatible 3D scaffold.

Monolayers of neurons and glia have been employed for decades as tools for the study of cellular physiology and as the basis for a variety of standard toxicological assays. A variety of three dimensional (3D) culture techniques have been developed with the aim to produce cultures that recapitulate desirable features of intact. In this study, we investigated the effect of preparing primary mouse mixed neuron and glial cultures in the inert 3D scaffold, Alvetex. Using planar multielectrode arrays, we compared the spontaneous bioelectrical activity exhibited by neuroglial networks grown in the scaffold with that seen in the same cells prepared as conventional monolayer cultures. Two dimensional (monolayer; 2D) cultures exhibited a significantly higher spike firing rate than that seen in 3D cultures although no difference was seen in total signal power (<50Hz) while pharmacological responsiveness of each culture type to antagonism of GABAAR, NMDAR and AMPAR was highly comparable. Interestingly, correlation of burst events, spike firing and total signal power (<50Hz) revealed that local field potential events were associated with action potential driven bursts as was the case for 2D cultures. Moreover, glial morphology was more physiologically normal in 3D cultures. These results show that 3D culture in inert scaffolds represents a more physiologically normal preparation which has advantages for physiological, pharmacological, toxicological and drug development studies, particularly given the extensive use of such preparations in high throughput and high content systems.

Effects and mechanisms of ginseng and ginsenosides on cognition.

Reviewed here is the existing evidence for the effects of ginseng extracts and isolated ginsenosides relevant to cognition in humans. Clinical studies in healthy volunteers and in patients with neurological disease or deficit, evidence from preclinical models of cognition, and pharmacokinetic data are considered. Conditions under which disease modification may indirectly benefit cognition but may not translate to cognitive benefits in healthy subjects are discussed. The number of chronic studies of ginseng effects in healthy individuals is limited, and the results from acute studies are inconsistent, making overall assessment of ginseng's efficacy as a cognitive enhancer premature. However, mechanistic results are encouraging; in particular, the ginsenosides Rg3 , Rh1 , Rh2 , Rb1 , Rd, Rg2 , and Rb3 , along with the aglycones protopanaxadiol and protopanaxatriol, warrant further attention. Compound K has a promising pharmacokinetic profile and can affect neurotransmission and neuroprotection. Properly conducted trials using standardized tests in healthy individuals reflecting the target population for ginseng supplementation are required to address inconsistencies in results from acute studies. The evidence summarized here suggests ginseng has potential, but unproven, benefits on cognition.

Voltage-gated sodium (NaV) channel blockade by plant cannabinoids does not confer anticonvulsant effects per se.

Cannabidiol (CBD) is a non-psychoactive, well-tolerated, anticonvulsant plant cannabinoid, although its mechanism(s) of seizure suppression remains unknown. Here, we investigate the effect of CBD and the structurally similar cannabinoid, cannabigerol (CBG), on voltage-gated Na(+) (NaV) channels, a common anti-epileptic drug target. CBG's anticonvulsant potential was also assessed in vivo. CBD effects on NaV channels were investigated using patch-clamp recordings from rat CA1 hippocampal neurons in brain slices, human SH-SY5Y (neuroblastoma) cells and mouse cortical neurons in culture. CBG effects were also assessed in SH-SY5Y cells and mouse cortical neurons. CBD and CBG effects on veratridine-stimulated human recombinant NaV1.1, 1.2 or 1.5 channels were assessed using a membrane potential-sensitive fluorescent dye high-throughput assay. The effect of CBG on pentyleneterazole-induced (PTZ) seizures was assessed in rat. CBD (10µM) blocked NaV currents in SH-SY5Y cells, mouse cortical neurons and recombinant cell lines, and affected spike parameters in rat CA1 neurons; CBD also significantly decreased membrane resistance. CBG blocked NaV to a similar degree to CBD in both SH-SY5Y and mouse recordings, but had no effect (50-200mg/kg) on PTZ-induced seizures in rat. CBD and CBG are NaV channel blockers at micromolar concentrations in human and murine neurons and recombinant cells. In contrast to previous reports investigating CBD, CBG had no effect upon PTZ-induced seizures in rat, indicating that NaV blockade per se does not correlate with anticonvulsant effects.

Home cervical ripening with dinoprostone gel in nulliparous women with singleton pregnancies.

OBJECTIVE: To evaluate whether home cervical ripening is safe and results in shorter hospital stay. METHODS: This was a retrospective cohort study of women with singleton pregnancies having induction of labor for postmaturity at a single center between January 2007 and June 2010. Women were offered home cervical ripening with 1 mg dinoprostone gel if they were nulliparous, had uncomplicated singleton pregnancies, and the indication for induction was postmaturity. RESULTS: Nine hundred seven of 1,536 (59.1%) nulliparous women having induction of labor for postmaturity were eligible for home cervical ripening. The median number of hours at home was 11.76 hours (range 0-24.82 hours). There were no cases of birth outside of the hospital, uterine rupture, or significant neonatal morbidity or neonatal death related to home cervical ripening. Eighty-five (5.5%) women who underwent hospital cervical ripening because of maternal preference or social issues formed a hospital cervical ripening comparison group. There was no significant difference in the total number of hours before delivery spent in the hospital between the two groups (26.25; 95% confidence interval [CI] 25.27-27.23 in home cervical ripening group compared with 24.28; 95% CI 22.5-26.0 in the hospital group; P=.26). CONCLUSIONS: Clinical outcomes are comparable in nulliparous women who receive a single dose of dinoprostone gel for home cervical ripening compared with those who undergo hospital cervical ripening. However, preadmission home cervical ripening with 1 mg dinoprostone does not decrease the number of hours women spend in the hospital. LEVEL OF EVIDENCE: II.

Influence of the menstrual cycle, pregnancy, and preeclampsia on arterial stiffness.

Arterial stiffness and compliance are major predictors of adverse cardiovascular events and are influenced by female sex hormones, including estrogen and progesterone. The aim of this longitudinal study was to evaluate the effect of the menstrual cycle, normal pregnancy, and preeclampsia on central and systemic arterial stiffness. Ten healthy nulliparous women with regular menses were studied in the early and midfollicular, periovulatory, and luteal phases of a single menstrual cycle. Twenty-two primigravida pregnant women were studied throughout pregnancy at 16, 24, 32, and 37 weeks gestation and at 7 weeks postpartum. Fifteen primigravida women with preeclampsia were studied at diagnosis and 7 weeks postpartum. Augmentation index and carotid-radial and carotid-femoral pulse wave velocities were measured using applanation tonometry. Augmentation index fell during the luteal phase of the menstrual cycle (luteal phase versus periovulatory phase; P<0.05). In normal pregnancy, pulse wave velocity and augmentation index increased from 24 weeks over the third trimester (P