High-threshold mechanosensitive ion channels blocked by a novel conopeptide mediate pressure-evoked pain

Little is known about the molecular basis of somatosensory mechanotransduction in mammals. We screened a library of peptide toxins for effects on mechanically activated currents in cultured dorsal root ganglion neurons. One conopeptide analogue, termed NMB-1 for noxious mechanosensation blocker 1, selectively inhibits (IC50 1 µM) sustained mechanically activated currents in a subset of sensory neurons. Biotinylated NMB-1 retains activity and binds selectively to peripherin-positive nociceptive sensory neurons. The selectivity of NMB-1 was confirmed by the fact that it has no inhibitory effects on voltage-gated sodium and calcium channels, or ligand-gated channels such as acid-sensing ion channels or TRPA1 channels. Conversely, the tarantula toxin, GsMTx-4, which inhibits stretch-activated ion channels, had no effects on mechanically activated currents in sensory neurons. In behavioral assays, NMB-1 inhibits responses only to high intensity, painful mechanical stimulation and has no effects on low intensity mechanical stimulation or thermosensation. Unexpectedly, NMB-1 was found to also be an inhibitor of rapid FM1-43 loading (a measure of mechanotransduction) in cochlear hair cells. These data demonstrate that pharmacologically distinct channels respond to distinct types of mechanical stimuli and suggest that mechanically activated sustained currents underlie noxious mechanosensation. NMB-1 thus provides a novel diagnostic tool for the molecular definition of channels involved in hearing and pressure-evoked pain

Life-history and diet of two populations of Natrix maura (Reptilia, Colubridae) from contrasted habitats in Sardinia

Aspects of general ecology and population biology of two populations of freeliving viperine snakes (Natrix maura) were studied in Mediterranean localities of the island of Sardinia (Thyrrenian Sea, Italy). The apparent adult sex-ratio was not significantly different from equality, but males were slightly more numerous than females. The diet consisted mainly of arnphibians, both adults and tadpoles (area A) or fish (area B). Females were significantly longer than males. Reproduction was yearly, and clutch size and female total length were positively correlated. A hundred and seven live offsprings were obtained from 135 eggs (79.2% of successful incubation rates). The proportion of unsuccessful incubation was not significantly correlated with either female total length or clutch size. There was no evidence of a trade-off between clutch size and offspring size, as hatchlings were no smaller in clutches that were unusually large in relation to maternal body size. Oviposition date was correlated with neither maternal length nor clutch size. lncubation period was significantly correlated with mean hatchling size, but not with either clutch size or maternal length. No female died after oviposition, thus suggesting a very low incidence of mortality rates dueto starvation and reproductive costs. A case of long-term sperm storage deriving from autumnal mating is described. Population size estimates are provided for both study areas. Key words: Natrix maura, Ecology, Mediterranean region, Sardinia.Aspects of general ecology and population biology of two populations of freeliving viperine snakes (Natrix maura) were studied in Mediterranean localities of the island of Sardinia (Thyrrenian Sea, Italy). The apparent adult sex-ratio was not significantly different from equality, but males were slightly more numerous than females. The diet consisted mainly of arnphibians, both adults and tadpoles (area A) or fish (area B). Females were significantly longer than males. Reproduction was yearly, and clutch size and female total length were positively correlated. A hundred and seven live offsprings were obtained from 135 eggs (79.2% of successful incubation rates). The proportion of unsuccessful incubation was not significantly correlated with either female total length or clutch size. There was no evidence of a trade-off between clutch size and offspring size, as hatchlings were no smaller in clutches that were unusually large in relation to maternal body size. Oviposition date was correlated with neither maternal length nor clutch size. lncubation period was significantly correlated with mean hatchling size, but not with either clutch size or maternal length. No female died after oviposition, thus suggesting a very low incidence of mortality rates dueto starvation and reproductive costs. A case of long-term sperm storage deriving from autumnal mating is described. Population size estimates are provided for both study areas. Key words: Natrix maura, Ecology, Mediterranean region, Sardinia.Aspects of general ecology and population biology of two populations of freeliving viperine snakes (Natrix maura) were studied in Mediterranean localities of the island of Sardinia (Thyrrenian Sea, Italy). The apparent adult sex-ratio was not significantly different from equality, but males were slightly more numerous than females. The diet consisted mainly of arnphibians, both adults and tadpoles (area A) or fish (area B). Females were significantly longer than males. Reproduction was yearly, and clutch size and female total length were positively correlated. A hundred and seven live offsprings were obtained from 135 eggs (79.2% of successful incubation rates). The proportion of unsuccessful incubation was not significantly correlated with either female total length or clutch size. There was no evidence of a trade-off between clutch size and offspring size, as hatchlings were no smaller in clutches that were unusually large in relation to maternal body size. Oviposition date was correlated with neither maternal length nor clutch size. lncubation period was significantly correlated with mean hatchling size, but not with either clutch size or maternal length. No female died after oviposition, thus suggesting a very low incidence of mortality rates dueto starvation and reproductive costs. A case of long-term sperm storage deriving from autumnal mating is described. Population size estimates are provided for both study areas. Key words: Natrix maura, Ecology, Mediterranean region, Sardinia

NMDA receptor subunit expression and PAR2 receptor activation in colospinal afferent neurons (CANs) during inflammation induced visceral hypersensitivity

<p>Abstract</p> <p>Background</p> <p>Visceral hypersensitivity is a clinical observation made when diagnosing patients with functional bowel disorders. The cause of visceral hypersensitivity is unknown but is thought to be attributed to inflammation. Previously we demonstrated that a unique set of enteric neurons, colospinal afferent neurons (CANs), co-localize with the NR1 and NR2D subunits of the NMDA receptor as well as with the PAR2 receptor. The aim of this study was to determine if NMDA and PAR2 receptors expressed on CANs contribute to visceral hypersensitivity following inflammation. Recently, work has suggested that dorsal root ganglion (DRG) neurons expressing the transient receptor potential vanilloid-1 (TRPV1) receptor mediate inflammation induced visceral hypersensitivity. Therefore, in order to study CAN involvement in visceral hypersensitivity, DRG neurons expressing the TRPV1 receptor were lesioned with resiniferatoxin (RTX) prior to inflammation and behavioural testing.</p> <p>Results</p> <p>CANs do not express the TRPV1 receptor; therefore, they survive following RTX injection. RTX treatment resulted in a significant decrease in TRPV1 expressing neurons in the colon and immunohistochemical analysis revealed no change in peptide or receptor expression in CANs following RTX lesioning as compared to control data. Behavioral studies determined that both inflamed non-RTX and RTX animals showed a decrease in balloon pressure threshold as compared to controls. Immunohistochemical analysis demonstrated that the NR1 cassettes, N1 and C1, of the NMDA receptor on CANs were up-regulated following inflammation. Furthermore, inflammation resulted in the activation of the PAR2 receptors expressed on CANs.</p> <p>Conclusion</p> <p>Our data show that inflammation causes an up-regulation of the NMDA receptor and the activation of the PAR2 receptor expressed on CANs. These changes are associated with a decrease in balloon pressure in response to colorectal distension in non-RTX and RTX lesioned animals. Therefore, these data suggest that CANs contribute to visceral hypersensitivity during inflammation.</p

Testing hypotheses of habitat use and temporal activity in relation to body plan in a mediterranean lizard community

A body plan (bauplan) is a suite of morphological characters shared by phylogenetically related animals at some point during their development. Despite its value, the bauplan concept is still rarely employed to characterize functional groups in community ecology. Here, we examine habitat use and spatio-temporal activity correlates of an entire sevenspecies community of lizards with different bauplans. The study was carried out in three locations in central Italy, encompassing a complex landscape with a patchy mosaic of a wide variety of habitats and microclimates. We tested four hypotheses regarding niche breadth, habitat use, and activity patterns. The first hypothesis, niche complementarity, in which species with similar body shapes should non-randomly partition available habitats, was not supported. By contrast, the hypotheses that larger bodied species should have a wider niche breadth, that slower species should inhabit habitat types of higher cover, and species inhabiting open sunny habitats should exhibit more seasonally variable activity patterns, were all supported by the data. Sympatric lizard communities in our study area were clearly organized by autecological constraints and eco-physiological attributes

Absence of lenadogene nolparvovec DNA in a brain tumor biopsy from a patient in the REVERSE clinical study, a case report

Background: Leber Hereditary Optic Neuropathy (LHON) is a rare, maternally-inherited mitochondrial disease that primarily affects retinal ganglion cells (RGCs) and their axons in the optic nerve, leading to irreversible, bilateral severe vision loss. Lenadogene nolparvovec gene therapy was developed as a treatment for patients with vision loss from LHON caused by the most prevalent m.11778G &gt; A mitochondrial DNA point mutation in the MT-ND4 gene. Lenadogene nolparvovec is a replication-defective recombinant adeno-associated virus vector 2 serotype 2 (AAV2/2), encoding the human wild-type MT-ND4 protein. Lenadogene nolparvovec was administered by intravitreal injection (IVT) in LHON patients harboring the m.11778G &gt; A ND4 mutation in a clinical development program including one phase 1/2 study (REVEAL), three phase 3 pivotal studies (REVERSE, RESCUE, REFLECT), and one long-term follow-up study (RESTORE, the follow-up of REVERSE and RESCUE patients). Case presentation: A 67-year-old woman with MT-ND4 LHON, included in the REVERSE clinical study, received a unilateral IVT of lenadogene nolparvovec in the right eye and a sham injection in the left eye in May 2016, 11.4 months and 8.8 months after vision loss in her right and left eyes, respectively. The patient had a normal brain magnetic resonance imaging with contrast at the time of diagnosis of LHON. Two years after treatment administration, BCVA had improved in both eyes. The product was well tolerated with mild and resolutive anterior chamber inflammation in the treated eye. In May 2019, the patient was diagnosed with a right temporal lobe glioblastoma, IDH-wildtype, World Health Organization grade 4, based on histological analysis of a tumor excision. The brain tumor was assessed for the presence of vector DNA by using a sensitive validated qPCR assay targeting the ND4 sequence of the vector. Conclusion: ND4 DNA was not detected (below 15.625 copies/μg of genomic DNA) in DNA extracted from the brain tumor, while a housekeeping gene DNA was detected at high levels. Taken together, this data shows the absence of detection of lenadogene nolparvovec in a brain tumor (glioblastoma) of a treated patient in the REVERSE clinical trial 3 years after gene therapy administration, supporting the long-term favorable safety of lenadogene nolparvovec

TRPA1 Mediates Mechanical Currents in the Plasma Membrane of Mouse Sensory Neurons

Mechanosensitive channels serve as essential sensors for cells to interact with their environment. The identity of mechanosensitive channels that underlie somatosensory touch transduction is still a mystery. One promising mechanotransduction candidate is the Transient Receptor Potential Ankyrin 1 (TRPA1) ion channel. To determine the role of TRPA1 in the generation of mechanically-sensitive currents, we used dorsal root ganglion (DRG) neuron cultures from adult mice and applied rapid focal mechanical stimulation (indentation) to the soma membrane. Small neurons (diameter <27 µm) were studied because TRPA1 is functionally present in these neurons which largely give rise to C-fiber afferents in vivo. Small neurons were classified by isolectin B4 binding

Painful and painless mutations of SCN9A and SCN11A voltage-gated sodium channels

Chronic pain is a global problem affecting up to 20% of the world’s population and has a significant economic, social and personal cost to society. Sensory neurons of the dorsal root ganglia (DRG) detect noxious stimuli and transmit this sensory information to regions of the central nervous system (CNS) where activity is perceived as pain. DRG neurons express multiple voltage-gated sodium channels that underlie their excitability. Research over the last 20 years has provided valuable insights into the critical roles that two channels, NaV1.7 and NaV1.9, play in pain signalling in man. Gain of function mutations in NaV1.7 cause painful conditions while loss of function mutations cause complete insensitivity to pain. Only gain of function mutations have been reported for NaV1.9. However, while most NaV1.9 mutations lead to painful conditions, a few are reported to cause insensitivity to pain. The critical roles these channels play in pain along with their low expression in the CNS and heart muscle suggest they are valid targets for novel analgesic drugs