Nicolas Doyon

Morphine hyperalgesia gated through microglia-mediated disruption of neuronal Cl- homeostasis

A major unresolved issue in treating pain is the paradoxical hyperalgesia produced by the gold-standard analgesic morphine and other opiates. We found that hyperalgesia-inducing treatment with morphine resulted in downregulation of the K+-Cl− co-transporter KCC2, impairing Cl− homeostasis in rat spinal lamina l neurons. Restoring the anion equilibrium potential reversed the morphine-induced hyperalgesia without affecting tolerance. The hyperalgesia was also reversed by ablating spinal microglia. Morphine hyperalgesia, but not tolerance, required μ opioid receptor–dependent expression of P2X4 receptors (P2X4Rs) in microglia and μ-independent gating of the release of brain-derived neurotrophic factor (BDNF) by P2X4Rs. Blocking BDNF-TrkB signaling preserved Cl− homeostasis and reversed the hyperalgesia. Gene-targeted mice in which Bdnf was deleted from microglia did not develop hyperalgesia to morphine. However, neither morphine antinociception nor tolerance was affected in these mice. Our findings dissociate morphine-induced hyperalgesia from tolerance and suggest the microglia-to-neuron P2X4-BDNF-KCC2 pathway as a therapeutic target for preventing hyperalgesia without affecting morphine analgesia.

Efficacy of Synaptic Inhibition Depends on Multiple, Dynamically Interacting Mechanisms Implicated in Chloride Homeostasis

Chloride homeostasis is a critical determinant of the strength and robustness of inhibition mediated by GABAA receptors (GABAARs). The impact of changes in steady state Cl− gradient is relatively straightforward to understand, but how dynamic interplay between Cl− influx, diffusion, extrusion and interaction with other ion species affects synaptic signaling remains uncertain. Here we used electrodiffusion modeling to investigate the nonlinear interactions between these processes. Results demonstrate that diffusion is crucial for redistributing intracellular Cl− load on a fast time scale, whereas Cl−extrusion controls steady state levels. Interaction between diffusion and extrusion can result in a somato-dendritic Cl− gradient even when KCC2 is distributed uniformly across the cell. Reducing KCC2 activity led to decreased efficacy of GABAAR-mediated inhibition, but increasing GABAAR input failed to fully compensate for this form of disinhibition because of activity-dependent accumulation of Cl−. Furthermore, if spiking persisted despite the presence of GABAAR input, Cl− accumulation became accelerated because of the large Cl− driving force that occurs during spikes. The resulting positive feedback loop caused catastrophic failure of inhibition. Simulations also revealed other feedback loops, such as competition between Cl− and pH regulation. Several model predictions were tested and confirmed by [Cl−]i imaging experiments. Our study has thus uncovered how Cl− regulation depends on a multiplicity of dynamically interacting mechanisms. Furthermore, the model revealed that enhancing KCC2 activity beyond normal levels did not negatively impact firing frequency or cause overt extracellular K− accumulation, demonstrating that enhancing KCC2 activity is a valid strategy for therapeutic intervention.

Inhibitory synaptic plasticity: spike timing-dependence and putative network function

While the plasticity of excitatory synaptic connections in the brain has been widely studied, the plasticity of inhibitory connections is much less understood. Here, we present recent experimental and theoretical findings concerning the rules of spike timing-dependent inhibitory plasticity and their putative network function. This is a summary of a workshop at the COSYNE conference 2012.

Chloride Regulation: A Dynamic Equilibrium Crucial for Synaptic Inhibition.

Fast synaptic inhibition relies on tight regulation of intracellular Cl(-). Chloride dysregulation is implicated in several neurological and psychiatric disorders. Beyond mere disinhibition, the consequences of Cl(-) dysregulation are multifaceted and best understood in terms of a dynamical system involving complex interactions between multiple processes operating on many spatiotemporal scales. This dynamical perspective helps explain many unintuitive manifestations of Cl(-) dysregulation. Here we discuss how taking into account dynamical regulation of intracellular Cl(-) is important for understanding how synaptic inhibition fails, how to best detect that failure, why Cl(-) regulation is energetically so expensive, and the overall consequences for therapeutics.

Treating pathological pain: is KCC2 the key to the gate?

Despite intensive efforts, development of novel treatments of neuropathic pain has encountered mitigated success. While common and incapacitating, neuropathic pain remains poorly treated with few patients experiencing greater than 50% symptom relief [1]. As an example, pregabalin, which targets the α 2 δ-subunit of the voltage-dependent calcium channel and is the current gold standard for neuropathic pain, reduces the pain score significantly in only one out of four patients [2]. Targeting the TrpV1 channel, which has attracted considerable resources, has only yielded the weak-selling capsaicin patch and has recently been denied indication expansion into HIV-related neuropathic pain by the US FDA [101]. In addition to weak efficacy issues, most approved treatments cause common and significant central side effects such as sedation and dizziness, diminishing the quality of life of affected patients. Opiate-based treatments, for example, are plagued with several side effects beyond sedation, including psycho motor issues, abuse liability, tolerance and even paradoxical hyperalgesia [3]. Recent promising alternatives such as anti-NGF antibodies have demonstrated very strong analgesic efficacy, yet, these strategies were put on hold due to severe side effects; many patients developed joint damage [4]. The unmet needs of improved efficacy and a better adverse event profile requires a different approach. One potential way of treating neuropathic pain is to restore proper inhibitory pathways in the CNS. Indeed, central disinhibition has long been known to play a major role in the pathophysiology of neuropathic pain states [5]. After nerve injury, there is a marked decrease in the inhibitory efficacy of GABA A and glycine receptor-mediated transmission, which results in aberrant transmission of sensory and nociceptive information to the brain [6]. A first step to develop therapeutics that would restore proper inhibition is to understand the underlying mechanism of disinhibition. While several causes have been proposed [6], a key mechanism that has emerged involves the disruption of Cl homeostasis resulting from loss of activity of the K–Cl cotransporter KCC2, a molecule responsible for maintaining efficient Cl-mediated inhibition in central neurons [7]. This mechanism appears across several pathological pain syndromes with diverse etiologies, including spinal cord injury [8], inflammation [9], painful diabetic neuropathy [10], trigeminal pain [11] and more recently even morphine-induced paradoxical hyperalgesia [3]. Several components of the cellular signaling cascade underlying impaired Cl homeostasis in spinal nociceptive pathways in pathological conditions have been identified, uncovering several potential therapeutic targets [3,12]. Enhanced BDNF release either from microglia [3,12,13] or from sensory nerves [14], acting on TrkB receptors to cause downregulation of KCC2, appears as a common culprit in several conditions. In Treating pathological pain: is KCC2 the key to the gate?

Allosteric modulation of metabotropic glutamate receptors by chloride ions

Metabotropic glutamate receptors (mGluRs) play key roles in the modulation of many synapses. Chloride (Cl‐) is known to directly bind and regulate the function of different actors of neuronal activity, and several studies have pointed to the possible modulation of mGluRs by Cl‐. Herein, we demonstrate that Cl‐ behaves as a positive allosteric modulator of mGluRs. For example, whereas glutamate potency was 3.08 ± 0.33 μM on metabotropic glutamate (mGlu) 4 receptors in high‐Cl‐ buffer, signaling activity was almost abolished in low Cl_ in cell‐based assays. Cl‐ potency was 78.6 ± 3.5 mM. Cl‐ possesses a high positive cooperativity with glutamate (Hill slope ã6 on mGlu4), meaning that small variations in [Cl‐] lead to large variations in glutamate action. Using molecular modeling and mutagenesis, we have identified 2 well‐conserved Cl‐binding pockets in the extracellular domain of mGluRs. Moreover, modeling of activity‐dependent Cl‐ variations at GABAergic synapses suggests that these variations may be compatible with a dynamic modulation of the most sensitive mGluRs present in these synapses. Taken together, these data reveal a necessary role of Cl‐ for the glutamate activation of manymGluRs. Exploiting Cl‐ binding pockets may yield to the development of innovative regulators of mGluR activity.—Tora, A. S., Rovira, X., Dione, I., Bertrand, H.‐O., Brabet, I., De Koninck, Y., Doyon, N., Pin, J.P., Acher, F., Goudet, C. Allosteric modulation of metabotropic glutamate receptors by chloride ions. FASEB J. 29, 4174‐4188 (2015). www.fasebj.org

Mild KCC2 Hypofunction Causes Inconspicuous Chloride Dysregulation that Degrades Neural Coding

Disinhibition caused by Cl− dysregulation is implicated in several neurological disorders. This form of disinhibition, which stems primarily from impaired Cl− extrusion through the co-transporter KCC2, is typically identified by a depolarizing shift in GABA reversal potential (EGABA). Here we show, using computer simulations, that intracellular [Cl−] exhibits exaggerated fluctuations during transient Cl− loads and recovers more slowly to baseline when KCC2 level is even modestly reduced. Using information theory and signal detection theory, we show that increased Cl− lability and settling time degrade neural coding. Importantly, these deleterious effects manifest after less KCC2 reduction than needed to produce the gross changes in EGABA required for detection by most experiments, which assess KCC2 function under weak Cl− load conditions. By demonstrating the existence and functional consequences of “occult” Cl− dysregulation, these results suggest that modest KCC2 hypofunction plays a greater role in neurological disorders than previously believed.

On the Distance Between Smooth Numbers

Abstract Let P(n) stand for the largest prime factor of n ≥ 2 and set P(1) = 1. For each integer n ≥ 2, let δ(n) be the distance to the nearest P(n)-smooth number, that is, to the nearest integer whose largest prime factor is no larger than that of n. We provide a heuristic argument showing that Σ n≤x 1/δ(n) = (4 log 2 – 2 + o(1))x as x → ∞. Moreover, given an arbitrary real-valued arithmetic function ƒ, we study the behavior of the more general function δ ƒ(n) defined by δ ƒ(n) = min1≤m≠n, ƒ(m)≤ƒ(n) |n – m| for n ≥ 2, and δ ƒ (1) = 1. In particular, given any positive integers a < b, we show that Σ a≤n<b 1/δ ƒ(n) ≥ 2(b – a)/3 and that if ƒ(n) ≥ ƒ(a) for all n ∈ [a, b[, then one has Σ a<n<b δ ƒ(n) ≤ (b – a) log(b – a)/(2 log 2).

Improved Simulation of Electrodiffusion in the Node of Ranvier by Mesh Adaptation.

In neural structures with complex geometries, numerical resolution of the Poisson-Nernst-Planck (PNP) equations is necessary to accurately model electrodiffusion. This formalism allows one to describe ionic concentrations and the electric field (even away from the membrane) with arbitrary spatial and temporal resolution which is impossible to achieve with models relying on cable theory. However, solving the PNP equations on complex geometries involves handling intricate numerical difficulties related either to the spatial discretization, temporal discretization or the resolution of the linearized systems, often requiring large computational resources which have limited the use of this approach. In the present paper, we investigate the best ways to use the finite elements method (FEM) to solve the PNP equations on domains with discontinuous properties (such as occur at the membrane-cytoplasm interface). 1) Using a simple 2D geometry to allow comparison with analytical solution, we show that mesh adaptation is a very (if not the most) efficient way to obtain accurate solutions while limiting the computational efforts, 2) We use mesh adaptation in a 3D model of a node of Ranvier to reveal details of the solution which are nearly impossible to resolve with other modelling techniques. For instance, we exhibit a non linear distribution of the electric potential within the membrane due to the non uniform width of the myelin and investigate its impact on the spatial profile of the electric field in the Debye layer.

ON A THIN SET OF INTEGERS INVOLVING THE LARGEST PRIME FACTOR FUNCTION

For each integer n≥2, let P(n) denote its largest prime factor. Let S:={n≥2:n does not divide P(n)!} and S(x):=#{n≤x:n∈S}. Erdős (1991) conjectured that S is a set of zero density. This was proved by Kastanas (1994) who established that S(x)=O(x/logx). Recently, Akbik (1999) proved that S(x)=O(x exp{−(1/4)logx}). In this paper, we show that S(x)=x exp{−(2