The Essential Oil of Citrus lumia Risso and Poit. 'Pyriformis' Shows Promising Antioxidant, Anti-Inflammatory, and Neuromodulatory Effects.

Citrus lumia Risso and Poit. 'Pyriformis' are horticultural varieties of Citrus lumia Risso. The fruit is very fragrant and pear-shaped, with a bitter juice, a floral flavor, and a very thick rind. The flavedo shows enlarged (0.74 × 1.16 mm), spherical and ellipsoidal secretory cavities containing the essential oil (EO), visible using light microscopy, and more evident using scanning electron microscopy. The GC-FID and GC-MS analyses of the EO showed a phytochemical profile characterized by the predominance of D-limonene (93.67%). The EO showed interesting antioxidant and anti-inflammatory activities (IC50 0.07-2.06 mg/mL), as evaluated by the in vitro cell-free enzymatic and non-enzymatic assays. To evaluate the effect on the neuronal functional activity, the embryonic cortical neuronal networks grown on multi-electrode array chips were exposed to non-cytotoxic concentrations of the EO (5-200 µg/mL). The spontaneous neuronal activity was recorded and the mean firing rate, mean burst rate, percentage of spikes in a burst, mean burst durations and inter-spike intervals within a burst parameter were calculated. The EO induced strong and concentration-dependent neuroinhibitory effects, with IC50 ranging between 11.4-31.1 µg/mL. Furthermore, it showed an acetylcholinesterase inhibitory activity (IC50 0.19 mg/mL), which is promising for controlling some of the key symptoms of neurodegenerative diseases such as memory and cognitive concerns.

Innovative Quantitative Assessment of Hand Function in Carpal Tunnel Syndrome.

BACKGROUND: Carpal tunnel syndrome (CTS) compromises fine sensorimotor function during activities of daily living and affects a large number of individuals with high burden costs for society. The purpose of this study was to quantitatively characterize fine movement skills in CTS patients preoperatively and at 1 month postoperatively by means of a sensor-engineered glove, in order to provide new insights for evaluative and finally therapeutic purposes. METHODS: Forty-one CTS patients and 41 age- and gender-matched healthy controls (HC) were analyzed by adopting the engineered glove Hand Test System (HTS), which previously demonstrated its reliability and sensitivity to detect hands dysfunction in several neurological diseases. A sub-group of 11 CTS subjects was re-tested 1 month after surgery. Three parameters-touch duration (TD), inter-tapping interval (ITI), and movement rate (MR)-were considered to characterize hand function. RESULTS: The affected hand of CTS patients generally showed worst finger opposition performances than HC. Comparing the dominant hand, all parameters were able to significantly discriminate CTS patients from HC. Considering the nondominant hand, the best performing parameter in discriminating CTS from HC was TD. The follow-up assessment at 1 month after surgery showed that considered parameters were able to monitor patients' recovery. In particular, the TD parameter recorded at the 3 different assigned task modalities resulted significantly enhanced. CONCLUSIONS: Results of this pilot study proved the validity of the parameters obtained through the sensor-engineered glove to assess objectively hand functional status and surgical outcomes in CTS.

Cannabis-like activity of Zornia latifolia Sm. detected in vitro on rat cortical neurons: major role of the flavone syzalterin.

Zornia latifolia is a plant suspected to possess psychoactive properties and marketed as a marijuana substitute under the name 'maconha brava'. In this study, the effects of fractions obtained from a 2-propanol extract of aerial portions of the plant were determined by multielectrode array (MEA) analyses on cultured networks of rat cortical neurons. Lipophilic (ZL_lipo, mainly containing flavonoid aglycones), and hydrophilic (ZL_hydro, mainly containing flavonoid glycosides) fractions were initially obtained from the raw extract. ZL_lipo significantly inhibited mean firing rate (MFR) and mean bursting rate (MBR) of MEA recordings, while ZL_hydro induced no inhibition. Column chromatography separation of ZL_lipo yielded five fractions (ZL1-ZL5), among which ZL1 induced the strongest MFR and MBR inhibition. NMR and HPLC-MS analyses of ZL1 revealed the prevalence of the common flavonoids genistein (1) and apigenin (2) (in about a 1:1 ratio), and the presence of the rare flavone syzalterin (6,8-dimethylapigenin) (3) as a minor compound. Exposures of MEA to apigenin and genistein standards did not induce the MFR and MBR inhibition observed with ZL1, whereas exposure to syzalterin standard or to a 1:9 mixture syzalterin-genistein induced effects similar to ZL1. These inhibitory effects were comparable to that observed with high-THC hashish, possibly accounting for the plant psychoactive properties. Data indicate that Z. latifolia, currently marketed as a free herbal product, should be subjected to measures of control. In addition, syzalterin showed distinctive pharmacological properties, opening the way to its possible exploitation as a neuroactive drug.

Characteristics and Applications of Technology-Aided Hand Functional Assessment: A Systematic Review.

Technology-aided hand functional assessment has received considerable attention in recent years. Its applications are required to obtain objective, reliable, and sensitive methods for clinical decision making. This systematic review aims to investigate and discuss characteristics of technology-aided hand functional assessment and their applications, in terms of the adopted sensing technology, evaluation methods and purposes. Based on the shortcomings of current applications, and opportunities offered by emerging systems, this review aims to support the design and the translation to clinical practice of technology-aided hand functional assessment. To this end, a systematic literature search was led, according to recommended PRISMA guidelines, in PubMed and IEEE Xplore databases. The search yielded 208 records, resulting into 23 articles included in the study. Glove-based systems, instrumented objects and body-networked sensor systems appeared from the search, together with vision-based motion capture systems, end-effector, and exoskeleton systems. Inertial measurement unit (IMU) and force sensing resistor (FSR) resulted the sensing technologies most used for kinematic and kinetic analysis. A lack of standardization in system metrics and assessment methods emerged. Future studies that pertinently discuss the pathophysiological content and clinimetrics properties of new systems are required for leading technologies to clinical acceptance.

Comparative and Functional Screening of Three Species Traditionally used as Antidepressants: Valeriana officinalis L., Valeriana jatamansi Jones ex Roxb. and Nardostachys jatamansi (D.Don) DC.

The essential oils (EOs) of three Caprifoliaceae species, the Eurasiatic Valeriana officinalis (Vo), the Himalayan Valeriana jatamansi (Vj) and Nardostachys jatamansi (Nj), are traditionally used to treat neurological disorders. Roots/rhizomes micromorphology, DNA barcoding and EOs phytochemical characterization were carried out, while biological effects on the nervous system were assessed by acetylcholinesterase (AChE) inhibitory activity and microelectrode arrays (MEA). Nj showed the highest inhibitory activity on AChE (IC50 67.15 µg/mL) followed by Vo (IC50 127.30 µg/mL) and Vj (IC50 246.84 µg/mL). MEA analyses on rat cortical neurons, carried out by recording mean firing rate (MFR) and mean bursting rate (MBR), revealed stronger inhibition by Nj (IC50 18.8 and 11.1 µg/mL) and Vo (16.5 and 22.5 µg/mL), compared with Vj (68.5 and 89.3 µg/mL). These results could be related to different EO compositions, since sesquiterpenes and monoterpenes significantly contribute to the observed effects, but the presence of oxygenated compounds such as aldehydes and ketones is a discriminating factor in determining the order of potency. Our multidisciplinary approach represents an important tool to avoid the adulteration of herbal drugs and permits the evaluation of the effectiveness of EOs that could be used for a wide range of therapeutic applications.

Promising in vitro antioxidant, anti-acetylcholinesterase and neuroactive effects of essential oil from two non-psychotropic Cannabis sativa L. biotypes.

The aim of this study was to compare the micro-morphological features of two different non-drug Cannabis sativa L. biotypes (Chinese accession G-309 and one fibrante variety) and to evaluate the phytochemical profile as well as some biological properties of the essential oils (EOs) obtained by hydrodistillation of dried flowering tops. After a micro-morphological evaluation by scanning electron microscopy, the phytochemical composition was analysed by GC-FID and GC-MS analyses. Antioxidant and anti-acetylcholinesterase properties were investigated by several in vitro cell-free assays, while neuroactive effects were evaluated on mouse cortical neuronal as well as human iPS cell-derived central nervous system cells grown on MEA chips. Both EOs showed strong antioxidant properties mainly attributable to the high content of hydroxylated compounds as well as significant anti-acetylcholinesterase activities (IC50 74.64 and 57.31 µg/ml for Chinese accession and fibrante variety, respectively). Furthermore, they showed a concentration-dependent inhibition of spontaneous electrical activity of human and mouse neuronal networks, with the fibrante variety, which showed the best activity (MFR, IC50 0.71 and 10.60 µg/ml, respectively). The observed biological activities could be due to a synergic effect between terpenes and phytocannabinoids, although in vivo studies, which clarify the molecular mechanism, are still lacking.

Essential oil of Citrus lumia Risso: Phytochemical profile, antioxidant properties and activity on the central nervous system.

The use of essential oils (EOs) is known since long time in traditional medicine and aromatherapy for the management of various oxidative stress-related disorders and has been further increased recently for their neuroprotective and anti-aging potentials as well as for reducing anxiety and stress. The purpose of this work was to evaluate, for the first time, the chemical composition of Citrus lumia Risso EO and its antioxidant, anti-cholinesterase, and neuroactive properties by cell-free and cell-based assays. The EO has shown strong antioxidant and free radical scavenging properties, particularly in hydrogen atom transfer based assays (ß-carotene bleaching and ORAC, IC50 22 µg/mL and 46 µg/mL, respectively), that can be attributed to the high content of monoterpenes, especially d-Limonene (48.905%), and Linalool (18.245%). Furthermore, the EO has shown an interesting anti-acetylcholinesterase activity (IC50 258.25 µg/mL). Data from MTT analysis indicate that the cytotoxicity of EO, evaluated on L929 mouse fibroblasts, is very low, with an IC50 higher than 500 µg/mL at 48 h. Rat neuronal networks subjected to EO showed a concentration-dependent inhibition of spontaneous electrical activity. Results indicate that C. lumia EO could be an important source of natural antioxidants suggesting an important preventive role in the onset of oxidative stress-related pathologies.

Generation of a Functional Human Neural Network by NDM29 Overexpression in Neuroblastoma Cancer Cells.

Recent advances in life sciences suggest that human and rodent cell responses to stimuli might differ significantly. In this context, the results achieved in neurotoxicology and biomedical research practices using neural networks obtained from mouse or rat primary culture of neurons would benefit of the parallel evaluation of the same parameters using fully differentiated neurons with a human genetic background, thus emphasizing the current need of neuronal cells with human origin. In this work, we developed a human functionally active neural network derived by human neuroblastoma cancer cells genetically engineered to overexpress NDM29, a non-coding RNA whose increased synthesis causes the differentiation toward a neuronal phenotype. These cells are here analyzed accurately showing functional and morphological traits of neurons such as the expression of neuron-specific proteins and the possibility to generate the expected neuronal current traces and action potentials. Their morphometrical analysis is carried out by quantitative phase microscopy showing soma and axon sizes compatible with those of functional neurons. The ability of these cells to connect autonomously forming physical junctions recapitulates that of hippocampal neurons, as resulting by connect-ability test. Lastly, these cells self-organize in neural networks able to produce spontaneous firing, in which spikes can be clustered in bursts. Altogether, these results show that the neural network obtained by NDM29-dependent differentiation of neuroblastoma cells is a suitable tool for biomedical research practices.

Coriandrum sativum and Lavandula angustifolia Essential Oils: Chemical Composition and Activity on Central Nervous System.

The aims of this study are to determine the chemical composition of Lavandula angustifolia Mill. and Coriandrum sativum L. essential oils, to evaluate their cytotoxic effects in SH-SY5Y human neuroblastoma cells, to investigate whether an alteration of adenylate cyclase 1 (ADCY1) and of extracellular signal-regulated kinase (ERK) expression can take part in the molecular mechanisms of the essential oils, and to study their possible neuronal electrophysiological effects. The essential oils were obtained by hydrodistillation, and studied by GC and GC-MS. In the oils from L. angustifolia and C. sativum, linalool was the main component (33.1% and 67.8%, respectively). SH-SY5Y cells were incubated with different concentrations of essential oils and of linalool. Cell viability and effects on ADCY1 and ERK expression were analyzed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide MTT and Western blotting, respectively. Variation in cellular electrophysiology was studied in primary cultures of rat cortical neurons with a multi-electrode array (MEA)-based approach. The essential oils and linalool revealed different cytotoxic activities. Linalool inhibited ADCY1 and ERK expression. Neuronal networks subjected to L. angustifolia and C. sativum essential oils showed a concentration-dependent inhibition of spontaneous electrical activity.

Microelectrode array (MEA) platform as a sensitive tool to detect and evaluate Ostreopsis cf. ovata toxicity.

In the last decade, the occurrence of harmful dinoflagellate blooms of the genus Ostreopsis has increased both in frequency and in geographic distribution with adverse impacts on public health and the economy. Ostreopsis species are producers of palytoxin-like toxins (putative palytoxin and ovatoxins) which are among the most potent natural non-protein compounds known to date, exhibiting extreme toxicity in mammals, including humans. Most existing toxicological data are derived from in vivo mouse assay and are related to acute effects of pure palytoxin, without considering that the toxicity mechanism of dinoflagellates can be dependent on the varying composition of complex biotoxins mixture and on the presence of cellular components. In this study, in vitro neuronal networks coupled to microelectrode array (MEA)-based system are proposed, for the first time, as sensitive biosensors for the evaluation of marine alga toxicity on mammalian cells. Toxic effect was investigated by testing three different treatments of laboratory cultured Ostreopsis cf. ovata cells: filtered and re-suspended algal cells; filtered, re-suspended and sonicated algal cells; conditioned growth medium devoid of algal cells. The great sensitivity of this system revealed the mixture of PTLX-complex analogues naturally released in the growth medium and the different potency of the three treatments to inhibit the neuronal network spontaneous electrical activity. Moreover, by means of the multiparametric analysis of neuronal network activity, the approach revealed a different toxicity mechanism of the cellular component compared to the algal conditioned growth medium, highlighting the potential active role of the first treatment.

Calcium-permeable AMPA receptors trigger vesicular glutamate release from Bergmann gliosomes.

The Bergmann glia is equipped with Ca2+-permeable AMPA receptors for glutamate, indispensable for structural and functional relations between the Bergmann glia and parallel/climbing fibers-Purkinje cell synapses. To better understand roles for the Bergmann AMPA receptors, herein we investigate on gliotransmitter release and Ca2+ signals in isolated Bergmann glia processes obtained from adult rat cerebellum. We found that: 1) the rat cerebellar purified astrocyte processes (gliosomes) expressed astrocytic and Bergmann markers and exhibited negligible contamination by nerve terminals, microglia, or oligodendrocytes; 2) activation of Ca2+-permeable AMPA receptors caused Ca2+ signals in the processes, and the release of glutamate from the processes; 3) effectiveness of rose bengal, trypan blue or bafilomycin A1, indicated that activation of the AMPA receptors evoked vesicular glutamate release. Cerebellar purified nerve terminals appeared devoid of glutamate-releasing Ca2+-permeable AMPA receptors, indicating that neuronal contamination may not be the source of the signals detected. Ultrastructural analysis indicated the presence of vesicles in the cytoplasm of the processes; confocal imaging confirmed the presence of vesicular glutamate transporters in Bergmann glia processes. We conclude that: a vesicular mechanism for release of the gliotransmitter glutamate is present in mature Bergmann processes; entry of Ca2+ through the AMPA receptors located on Bergmann processes is coupled with vesicular glutamate release. The findings would add a new role for a well-known Bergmann target for glutamate (the Ca2+-permeable AMPA receptors) and a new actor (the gliotransmitter glutamate) at the cerebellar excitatory synapses onto Purkinje cells.

Multiparametric characterisation of neuronal network activity for in vitro agrochemical neurotoxicity assessment.

The last few decades have seen the marketing of hundreds of new pesticide products with a forecasted expansion of the global agrochemical industry. As several pesticides directly target nervous tissue as their mechanism of toxicity, alternative methods to routine in vivo animal testing, such as the Multi Electrode Array (MEAs)-based approach, have been proposed as an in vitro tool to perform sensitive, quick and low cost neuro-toxicological screening. Here, we examined the effects of a training set of eleven active substances known to have neuronal or non-neuronal targets, contained in the most commonly used agrochemicals, on the spontaneous electrical activity of cortical neuronal networks grown on MEAs. A multiparametric characterisation of neuronal network firing and bursting was performed with the aim of investigating how this can contribute to the efficient evaluation of in vitro chemical-induced neurotoxicity. The analysis of MFR, MBR, MBD, MISI_B and % Spikes_B parameters identified four different groups of chemicals: one wherein only inhibition is observed (chlorpyrifos, deltamethrin, orysastrobin, dimoxystrobin); a second one in which all parameters, except the MISI_B, are inhibited (carbaryl, quinmerac); a third in which increases at low chemical concentration are followed by decreases at high concentration, with exception of MISI_B that only decreased (fipronil); a fourth in which no effects are observed (paraquat, glyphosate, imidacloprid, mepiquat). The overall results demonstrated that the multiparametric description of the neuronal networks activity makes MEA-based screening platform an accurate and consistent tool for the evaluation of the toxic potential of chemicals. In particular, among the bursting parameters the MISI_B was the best that correlates with potency and may help to better define chemical toxicity when MFR is affected only at relatively high concentration.

The P2X7 receptor as a route for non-exocytotic glutamate release: dependence on the carboxyl tail.

P2X7 receptors trigger Ca(2+) -dependent exocytotic glutamate release, but also function as a route for non-exocytotic glutamate release from neurons or astrocytes. To gain an insight into the mechanisms involving the P2X7 receptor as a direct pathway for glutamate release, we compared the behavior of a full-length rat P2X7 receptor, a truncated rat P2X7 receptor in which the carboxyl tail had been deleted, a rat P2X7 receptor with the 18-amino acid cysteine-rich motif of the carboxyl tail deleted, and a rat P2X2 receptor, all of which are expressed in HEK293 cells. We found that the P2X7 receptor function as a route for glutamate release was antagonized in a non-competitive way by extracellular Mg(2+) , did not require the recruitment of pore-forming molecules, and was dependent on the carboxyl tail. Indeed, the truncated P2X7 receptor and the P2X7 receptor with the deleted cysteine-rich motif both lost their function as a pathway for glutamate release, while still evoking intracellular Ca(2+) elevation. No glutamate efflux was observed through the P2X2 receptor. Notably, HEK293 cells (lacking the machinery for Ca(2+) -dependent exocytosis), when transfected with P2X7 receptors, appear to be a suitable model for investigating the P2X7 receptor as a route for non-exocytotic glutamate efflux.

Calmidazolium selectively inhibits exocytotic glutamate release evoked by P2X7 receptor activation.

We previously observed that activation of presynaptic P2X7 receptors located on rat cerebrocortical nerve terminals induced the release of glutamate through different modes: the channel conformation allowing Ca(2+) entry triggered exocytotic release, while the receptor itself functioned as a permeation pathway for the non-exocytotic glutamate release. Considering that exocytotic and non-exocytotic glutamate release evoked by the activation of P2X7 receptors might play a role in the control of glutamatergic synapses, we investigated whether calmidazolium (which has been found to inhibit small cation currents through recombinant P2X7 receptors, but not organic molecule permeation) could distinguish between P2X7-related exocytotic and non-exocytotic modes of glutamate release. We found that calmidazolium inhibited the intrasynaptosomal Ca(2+) response to P2X7 receptor activation and the Ca(2+)-dependent exocytotic glutamate release from rat cerebrocortical nerve terminals, but was ineffective against the Ca(2+)-independent glutamate release. The P2X7 competitive antagonist A-438079 eliminated both exocytotic and non-exocytotic P2X7 receptor-evoked glutamate release. Selective inhibition of exocytotic glutamate release indicates that calmidazolium inhibits events dependent on the function of native rat P2X7 receptors as Ca(2+) channels, and suggests that it can be used as a tool to dissociate P2X7-evoked exocytotic from non-exocytotic glutamate release.

UDP exerts cytostatic and cytotoxic actions in human neuroblastoma SH-SY5Y cells over-expressing P2Y6 receptor.

The role of P2 receptors for purines/pyrimidines is not well characterized in neuroblastoma, although a variety of purinergic mRNAs/proteins are expressed in these cells. Among these, the P2Y(6) receptor is the only subtype distinguished by UDP-specific activation. In this work, after over-expressing the P2Y(6) protein in human neuroblastoma SH-SY5Y cells, we find that UDP arrests cell cycle and induces apoptosis, by counteracting the pathological functioning of neuroblastoma in vitro. UDP also causes mitochondrial damage through diffusion of cytochrome c in the cytoplasm, and stimulates caspase-3,7,8 activities, with extensive over-expression of manganese superoxide dismutase. Our data establish the direct toxic role and anti-cancer activity of UDP in a neuroblastoma cell line, and identify the P2Y(6) receptor as a novel potential target in anti-tumoural therapies. This constitutes an advancement not only in the knowledge of purinergic signalling, but also in the biological and pathological aspects of neuroblastoma in vitro.