Deranged calcium signaling and neurodegeneration in spinocerebellar ataxia type 3

Spinocerebellar ataxia type 3 (SCA3), also known as Machado-Joseph disease (MJD), is an autosomal-dominant neurodegenerative disorder caused by a polyglutamine expansion in ataxin-3 (SCA3, MJD1) protein. In biochemical experiments we demonstrate that mutant SCA3exp specifically associated with the type 1 inositol 1,4,5-trisphosphate receptor (InsP3R1), an intracellular calcium (Ca2+) release channel. In electrophysiological and Ca2+ imaging experiments we show that InsP3R1 are sensitized to activation by InsP3 in the presence of mutant SCA3exp. We found that feeding SCA3-YAC-84Q transgenic mice with dantrolene, a clinically relevant stabilizer of intracellular Ca2+ signaling, improved their motor performance and prevented neuronal cells loss in pontine nuclei and substantia nigra regions. Our results indicate that deranged Ca2+ signaling may play an important role in SCA3 pathology and that Ca2+ signaling stabilizers such as dantrolene may be considered as potential therapeutic drugs for treatment of SCA3 patients

Ontogeny of purinergic receptor-regulated Ca2+ signaling in mouse cortical collecting duct epithelium

Changes in ATP-induced increase in {[}Ca2+], during collecting duct ontogeny were studied in primary monolayer cultures of mouse ureteric bud (UB) and cortical collecting duct (CCD) cells by Fura-PE3 fluorescence ratio imaging. In UB (embryonic day E14 and postnatal day P1) the ATIP-stimulated increase (EC50 approximate to 1 muM) in fluorescence ratio (DeltaR(ATP)) was independent of extracellular Ca2+ and insensitive to the P2 purinoceptor-antagonist suramin (1 mM). From day P7 onward when CCD morphogenesis had been completed DeltaR(ATP) increased and became dependent on extracellular Ca2+. This ATP-stimulated Ca2+ entry into CCD cells was non-capacitative and suramin (11 mM)insensitive, but sensitive to nifedipine (30 muM) and enhanced by Bay K8644 (15 muM), a blocker and an agonist of L-type Ca2+ channels, respectively. Quantitative RT-PCR demonstrated similar mRNA expression of L-type Ca2+ channel alpha1-subunit, P2Y(1), P2Y(2), and P2X(4b) purinoceptors in UB and CCD monolayers while the abundance of P2X(4) mRNA increased with CCD morphogenesis. In conclusion, both embryonic and postnatal cells express probably P2Y(2)-stimulated Ca2+ release from intracellular stores. With development, the CCD epithelium acquires ATP-stimulated Ca2+ entry via L-type Ca2+ channels. This pathway might by mediated by the increasing expression of P2X(4)-receptors resulting in an increasing ATP-dependent membrane depolarization and activation of L-type Ca2+ channels. Copyright (C) 2002 S. Karger AG, Basel

Subcellular heterogeneity of ryanodine receptor properties in ventricular myocytes with low T-tubule density

Rationale: In ventricular myocytes of large mammals, not all ryanodine receptor (RyR) clusters are associated with T-tubules (TTs); this fraction increases with cellular remodeling after myocardial infarction (MI). Objective: To characterize RyR functional properties in relation to TT proximity, at baseline and after MI. Methods: Myocytes were isolated from left ventricle of healthy pigs (CTRL) or from the area adjacent to a myocardial infarction (MI). Ca2+ transients were measured under whole-cell voltage clamp during confocal linescan imaging (fluo-3) and segmented according to proximity of TTs (sites of early Ca2+ release, F>F50 within 20 ms) or their absence (delayed areas). Spontaneous Ca2+ release events during diastole, Ca2+ sparks, reflecting RyR activity and properties, were subsequently assigned to either category. Results: In CTRL, spark frequency was higher in proximity of TTs, but spark duration was significantly shorter. Block of Na+/Ca2+ exchanger (NCX) prolonged spark duration selectively near TTs, while block of Ca2+ influx via Ca2+ channels did not affect sparks properties. In MI, total spark mass was increased in line with higher SR Ca2+ content. Extremely long sparks (>47.6 ms) occurred more frequently. The fraction of near-TT sparks was reduced; frequency increased mainly in delayed sites. Increased duration was seen in near-TT sparks only; Ca2+ removal by NCX at the membrane was significantly lower in MI. Conclusion: TT proximity modulates RyR cluster properties resulting in intracellular heterogeneity of diastolic spark activity. Remodeling in the area adjacent to MI differentially affects these RyR subpopulations. Reduction of the number of sparks near TTs and reduced local NCX removal limit cellular Ca2+ loss and raise SR Ca2+ content, but may promote Ca2+ waves

Disentangling astroglial physiology with a realistic cell model in silico

Electrically non-excitable astroglia take up neurotransmitters, buffer extracellular K+ and generate Ca2+ signals that release molecular regulators of neural circuitry. The underlying machinery remains enigmatic, mainly because the sponge-like astrocyte morphology has been difficult to access experimentally or explore theoretically. Here, we systematically incorporate multi-scale, tri-dimensional astroglial architecture into a realistic multi-compartmental cell model, which we constrain by empirical tests and integrate into the NEURON computational biophysical environment. This approach is implemented as a flexible astrocyte-model builder ASTRO. As a proof-of-concept, we explore an in silico astrocyte to evaluate basic cell physiology features inaccessible experimentally. Our simulations suggest that currents generated by glutamate transporters or K+ channels have negligible distant effects on membrane voltage and that individual astrocytes can successfully handle extracellular K+ hotspots. We show how intracellular Ca2+ buffers affect Ca2+ waves and why the classical Ca2+ sparks-and-puffs mechanism is theoretically compatible with common readouts of astroglial Ca2+ imaging

Intravital imaging reveals p53-dependent cancer cell death induced by phototherapy via calcium signaling.

One challenge in biology is signal transduction monitoring in a physiological context. Intravital imaging techniques are revolutionizing our understanding of tumor and host cell behaviors in the tumor environment. However, these deep tissue imaging techniques have not yet been adopted to investigate the second messenger calcium (Ca2+). In the present study, we established conditions that allow the in vivo detection of Ca2+ signaling in three-dimensional tumor masses in mouse models. By combining intravital imaging and a skinfold chamber technique, we determined the ability of photodynamic cancer therapy to induce an increase in intracellular Ca2+ concentrations and, consequently, an increase in cell death in a p53-dependent pathway

When, where and how? Focus on neuronal calcium dysfunctions in Alzheimer's Disease.

Alzheimer\u2019s disease (AD), since its characterization as a precise form of dementia with its own pathological hallmarks, has captured scientists\u2019 attention because of its complexity. The last 30 years have been filled with discoveries regarding the elusive aetiology of this disease and, thanks to advances in molecular biology and live imaging techniques, we now know that an important role is played by calcium (Ca2+). Ca2+, as ubiquitous second messenger, regulates a vast variety of cellular processes, from neuronal excitation and communication, to muscle fibre contraction and hormone secretion, with its action spanning a temporal scale that goes from microseconds to hours. It is therefore very challenging to conceive a single hypothesis that can integrate the numerous findings on this issue with those coming from the classical fields of AD research such as amyloid-beta (A) and tau pathology. In this contribution, we will focus our attention on the Ca2+ hypothesis of AD, dissecting it, as much as possible, in its subcellular localization, where the Ca2+ signal meets its specificity. We will also follow the temporal evolution of the Ca2+ hypothesis, providing some of the most updated discoveries. Whenever possible, we will link the findings regarding Ca2+ dysfunction to the other players involved in AD pathogenesis, hoping to provide a crossover body of evidence, useful to amplify the knowledge that will lead towards the discovery of an effective therapy

Dorsal-CA1 hippocampal neuronal ensembles encode nicotine-reward contextual associations

Natural and drug rewards increase the motivational valence of stimuli in the environment that, through Pavlovian learning mechanisms, become conditioned stimuli that directly motivate behavior in the absence of the original unconditioned stimulus. While the hippocampus has received extensive attention for its role in learning and memory processes, less is known regarding its role in drug-reward associations. We used in vivo Ca2+ imaging in freely moving mice during the formation of nicotine preference behavior to examine the role of the dorsal-CA1 region of the hippocampus in encoding contextual reward-seeking behavior. We show the development of specific neuronal ensembles whose activity encodes nicotine-reward contextual memories and that are necessary for the expression of place preference. Our findings increase our understanding of CA1 hippocampal function in general and as it relates to reward processing by identifying a critical role for CA1 neuronal ensembles in nicotine place preference

Synaptic actions of amyotrophic-lateral-sclerosis-associated G85R-SOD1 in the squid giant synapse

© The Author(s), 2020. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Song, Y. Synaptic actions of amyotrophic-lateral-sclerosis-associated G85R-SOD1 in the squid giant synapse. Eneuro, (2020): ENEURO.0369-19.2020, doi: 10.1523/ENEURO.0369-19.2020.Altered synaptic function is thought to play a role in many neurodegenerative diseases, but little is known about the underlying mechanisms for synaptic dysfunction. The squid giant synapse (SGS) is a classical model for studying synaptic electrophysiology and ultrastructure, as well as molecular mechanisms of neurotransmission. Here, we conduct a multidisciplinary study of synaptic actions of misfolded human G85R-SOD1 causing familial Amyotrophic Lateral Sclerosis (fALS). G85R-SOD1, but not WT-SOD1, inhibited synaptic transmission, altered presynaptic ultrastructure, and reduced both the size of the Readily Releasable Pool (RRP) of synaptic vesicles and mobility from the Reserved Pool (RP) to the RRP. Unexpectedly, intermittent high frequency stimulation (iHFS) blocked inhibitory effects of G85R-SOD1 on synaptic transmission, suggesting aberrant Ca2+ signaling may underlie G85R-SOD1 toxicity. Ratiometric Ca2+ imaging showed significantly increased presynaptic Ca2+ induced by G85R-SOD1 that preceded synaptic dysfunction. Chelating Ca2+ using EGTA prevented synaptic inhibition by G85R-SOD1, confirming the role of aberrant Ca2+ in mediating G85R-SOD1 toxicity. These results extended earlier findings in mammalian motor neurons and advanced our understanding by providing possible molecular mechanisms and therapeutic targets for synaptic dysfunctions in ALS as well as a unique model for further studies.Grass Foundation, HHMI, MGH Jack Satter Foundation, Harvard University ALS and Alzheimer's Endowed Research Fund, Harvard Brain Science Initiative