A major unresolved issue in treating pain is the paradoxical hyperalgesia produced by the gold-standard analgesic morphine and other opiates. We show here that hyperalgesia-inducing treatment with morphine causes downregulation of the K+-Cl− cotransporter KCC2, impairing Cl− homeostasis in spinal lamina l neurons. Restoring Eanion 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 signalling preserved Cl− homeostasis and reversed the hyperalgesia. Gene-targeted mice in which BDNF was deleted from microglia did not develop hyperalgesia to morphine. Yet, neither morphine antinociception nor tolerance was affected in these animals. Our findings dissociate morphine-induced hyperalgesia from tolerance and unveil the microglia-to-neuron P2X4-BDNF-KCC2 pathway as a therapeutic target to prevent hyperalgesia without affecting morphine analgesia.

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Microglia-neuron interactions play a crucial role in several neurological disorders characterized by altered neural network excitability, such as epilepsy and neuropathic pain. While a series of potential messengers have been postulated as substrates of the communication between microglia and neurons, including cytokines, purines, prostaglandins, and nitric oxide, the specific links between messengers, microglia, neuronal networks, and diseases have remained elusive. Brain-derived neurotrophic factor (BDNF) released by microglia emerges as an exception in this riddle. Here, we review the current knowledge on the role played by microglial BDNF in controlling neuronal excitability by causing disinhibition. The efforts made by different laboratories during the last decade have collectively provided a robust mechanistic paradigm which elucidates the mechanisms involved in the synthesis and release of BDNF from microglia, the downstream TrkB-mediated signals in neurons, and the biophysical mechanism by which disinhibition occurs, via the downregulation of the K+-Cl− cotransporter KCC2, dysrupting Cl−homeostasis, and hence the strength of GABAA- and glycine receptor-mediated inhibition. The resulting altered network activity appears to explain several features of the associated pathologies. Targeting the molecular players involved in this canonical signaling pathway may lead to novel therapeutic approach for ameliorating a wide array of neural dysfunctions.

Neuropeptides are small protein molecules (composed of 3-100 amino-acid residues) that have been localized to discrete cell populations of central and peripheral neurons. In most instances, they coexist with low-molecular-weight neurotransmitters within the same neurons. At the subcellular level, neuropeptides are selectively stored, singularly or more frequently in combinations, within large granular vesicles. Release occurs through mechanisms different from classical calcium-dependent exocytosis at the synaptic cleft, and thus they account for slow synaptic and/or non-synaptic communication in neurons. Neuropeptide co-storage and coexistence can be observed throughout the central nervous system and are responsible for a series of functional interactions that occur at both pre- and post-synaptic levels. Thus, the subcellular site(s) of storage and sorting mechanisms into different neuronal compartments are crucial to the mode of release and the function of neuropeptides as neuronal messengers.

Ghrelin, an orexigenic peptide synthesized by endocrine cells of the gastric mucosa, is released in the bloodstream in response to a negative energetic status. Since discovery, the hypothalamus was identified as the main source of ghrelin in the CNS, and effects of the peptide have been mainly observed in this area of the brain. In recent years, an increasing number of studies have reported ghrelin synthesis and effects in specific populations of neurons also outside the hypothalamus. Thus, ghrelin activity has been described in midbrain, hindbrain, hippocampus, and spinal cord. The spectrum of functions and biological effects produced by the peptide on central neurons is remarkably wide and complex. It ranges from modulation of membrane excitability, to control of neurotransmitter release, neuronal gene expression, and neuronal survival and proliferation. There is not at present a general consensus concerning the source of ghrelin acting on central neurons. Whereas it is widely accepted that the hypothalamus represents the most important endogenous source of the hormone in CNS, the existence of extra-hypothalamic ghrelin-synthesizing neurons is still controversial. In addition, circulating ghrelin can theoretically be another natural ligand for central ghrelin receptors. This paper gives an overview on the distribution of ghrelin and its receptor across the CNS and critically analyses the data available so far as regarding the effects of ghrelin on central neurotransmission.

Brain-derived neurotrophic factor (BDNF) exerts its trophic effects by acting on the high-affinity specific receptor trkB. BDNF also modulates synaptic transmission in several areas of the CNS, including the spinal cord dorsal horn, where it acts as a pain modulator by yet incompletely understood mechanisms. Spinal neurons are the main source of trkB in lamina II (substantia gelatinosa). Expression of this receptor in dorsal root ganglion (DRG) cells has been a matter of debate, whereas a subpopulation of DRG neurons bears trkA receptors and contains BDNF. By the use of two different trkB antibodies we observed that 7.7% and 10.8% of DRG neurons co-expressed BDNF + trkB but not trkA, respectively, in rat and mouse. Ultrastructurally, full-length trkB (fl-trkB) receptors were present at somato-dendritic membranes of lamina II neurons (rat: 66.8%; mouse: 73.8%) and at axon terminals (rat: 33.2%; mouse: 26.2%). In both species, about 90% of these terminals were identified as primary afferent fibres (PAFs) considering their morphology and/or neuropeptide content. All fl-trkB-immunopositive C boutons in type Ib glomeruli were immunoreactive for BDNF and, at individual glomeruli and axo-dendritic synapses, fl-trkB receptors were located in a mutually exclusive fashion at pre- or postsynaptic membranes. Thus, only a small fraction of fl-trkB-immunoreactive dendrites were postsynaptic to BDNF-immunopositive PAFs. This is the first ultrastructural description of fl-trkB localization at synapses between first- and second-order sensory neurons in lamina II, and suggests that BDNF may be released by fl-trkB-immunopositive PAFs to modulate nociceptive input in this lamina of dorsal horn.

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GABA A /glycine-mediated neuronal inhibition critically depends on intracellular chloride (Cl −) concentration which is mainly regulated by the K +-Cl − co-transporter 2 (KCC2) in the adult central nervous system (CNS). KCC2 heterogeneity thus affects information processing across CNS areas. Here, we uncover a gradient in Cl − extrusion capacity across the superficial dorsal horn (SDH) of the spinal cord (laminae I-II: LI-LII), which remains concealed under low Cl − load. Under high Cl − load or heightened synaptic drive, lower Cl − extrusion is unveiled in LI, as expected from the gradient in KCC2 expression found across the SDH. Blocking TrkB receptors increases KCC2 in LI, pointing to differential constitutive TrkB activation across laminae. Higher Cl − lability in LI results in rapidly collapsing inhibition, and a form of activity-dependent synaptic plasticity expressed as a continuous facilitation of excitatory responses. The higher metaplasticity in LI as compared to LII differentially affects sensitization to thermal and mechanical input. Thus, inconspicuous heterogeneity of Cl − extrusion across laminae critically shapes plasticity for selective nociceptive modalities.