ATP Released by Injured Neurons Activates Schwann Cells

Injured nerve terminals of neuromuscular junctions (NMJs) can regenerate. This remarkable and complex response is governed by molecular signals that are exchanged among the cellular components of this synapse: motor axon nerve terminal (MAT), perisynaptic Schwann cells (PSCs), and muscle fiber. The nature of signals that govern MAT regeneration is ill-known. In the present study the spider toxin alpha-latrotoxin has been used as tool to investigate the mechanisms underlying peripheral neuroregeneration. Indeed this neurotoxin induces an acute, specific, localized and fully reversible damage of the presynaptic nerve terminal, and its action mimics the cascade of events that leads to nerve terminal degeneration in injured patients and in many neurodegenerative conditions. Here we provide evidence of an early release by degenerating neurons of adenosine triphosphate as alarm messenger, that contributes to the activation of a series of intracellular pathways within Schwann cells that are crucial for nerve regeneration: Ca2+, cAMP, ERK1/2, and CREB. These results contribute to define the cross-talk taking place among degenerating nerve terminals and PSCs, involved in the functional recovery of the NMJ

Mammalian gene expression variability is explained by underlying cell state.

Gene expression variability in mammalian systems plays an important role in physiological and pathophysiological conditions. This variability can come from differential regulation related to cell state (extrinsic) and allele-specific transcriptional bursting (intrinsic). Yet, the relative contribution of these two distinct sources is unknown. Here, we exploit the qualitative difference in the patterns of covariance between these two sources to quantify their relative contributions to expression variance in mammalian cells. Using multiplexed error robust RNA fluorescent in situ hybridization (MERFISH), we measured the multivariate gene expression distribution of 150 genes related to Ca2+ signaling coupled with the dynamic Ca2+ response of live cells to ATP. We show that after controlling for cellular phenotypic states such as size, cell cycle stage, and Ca2+ response to ATP, the remaining variability is effectively at the Poisson limit for most genes. These findings demonstrate that the majority of expression variability results from cell state differences and that the contribution of transcriptional bursting is relatively minimal

Studies of intercellular Ca2+ signaling and gap-junction coupling in the developing cochlea of mouse models affected by congenital hearing loss

Connexin 26 (Cx26) and connexin 30 (Cx30) form gap junction channels that allow the intercellular diffusion of the Ca2+ mobilizing second messenger IP3. They also form hemichannels that release ATP from the endolymphatic surface of cochlear supporting and epithelial cells. Released ATP in turn activates G-protein coupled P2Y2 and P2Y4 receptors, PLC-dependent generation of IP3, release of Ca2+ from intracellular stores, permitting the regenerative propagation of intercellular Ca2+ signals. In the course of this work, we found that cochlear non-sensory cells of the greater and lesser epithelial ridge (GER and LER, respectively) share the same PLC- and IP3R-dependent signal transduction cascade activated by ATP. In addition, we demonstrated that ATP-dependent Ca2+ signaling activity in cochlear non-sensory cells is spatially graded from the apex to the base of the cochlea during the first postnatal week. Ca2+ signaling under these conditions depends on inositol-1,4,5-trisphosphate generation from phospholipase C (PLC)-dependent hydrolysis of PI(4,5)P(2). Thus we analyzed mice with defective expression of PIPKIγ and found that (i) this enzyme is essential for the acquisition of hearing; (ii) it is primarily responsible for the synthesis of the receptor-regulated PLC-sensitive PI(4,5)P(2) pool in the cell syncytia that supports auditory hair cells and; (iii) spatially graded impairment of the PIP2-IP3-Ca2+ signaling pathway in cochlear non-sensory cells affects the level of gap junction coupling. Vice versa, we found defective gap junction coupling and intercellular IP3-dependent Ca2+ signaling the cochlea of mice with targeted ablation Cx26 or Cx30, as well as in mice knock in for a point mutation (Cx30T5M) associated with human congenital deafness. Altogether, our findings link bidirectionally defective hearing acquisition to Ca2+ signaling impairment and decreased biochemical coupling in the developing cochlea. Transduction of connexin deficient cochlear cultures with a bovine adeno associated virus vectors encoding Cx26 or Cx30 restored protein expression, rescued both gap junction coupling and Ca2+ signaling. Based on this work, we conclude that in vivo connexin gene delivery to the inner ear is a route worth exploring to rescue hearing function in mouse models of deafness and, in future, may lead to the development of therapeutic interventions in humans

Mammalian Brain As a Network of Networks

Acknowledgements AZ, SG and AL acknowledge support from the Russian Science Foundation (16-12-00077). Authors thank T. Kuznetsova for Fig. 6.Peer reviewedPublisher PD