Modulation of mechanosensory vibrissal responses in the trigeminocervical complex by stimulation of the greater occipital nerve in a rat model of trigeminal neuropathic pain.

Background Stimulation of the occipital or trigeminal nerves has been successfully used to treat chronic refractory neurovascular headaches such as migraine or cluster headache, and painful neuropathies. Convergence of trigeminal and occipital sensory afferents in the ‘trigeminocervical complex’ (TCC) from cutaneous, muscular, dural, and visceral sources is a key mechanism for the input-induced central sensitization that may underlie the altered nociception. Both excitatory (glutamatergic) and inhibitory (GABAergic and glycinergic) mechanisms are involved in modulating nociception in the spinal and medullary dorsal horn neurons, but the mechanisms by which nerve stimulation effects occur are unclear. This study was aimed at investigating the acute effects of electrical stimulation of the greater occipital nerve (GON) on the responses of neurons in the TCC to the mechanical stimulation of the vibrissal pad. Methods Adult male Wistar rats were used. Neuronal recordings were obtained in laminae II-IV in the TCC in control, sham and infraorbital chronic constriction injury (CCI-IoN) animals. The GON was isolated and electrically stimulated. Responses to the stimulation of vibrissae by brief air pulses were analyzed before and after GON stimulation. In order to understand the role of the neurotransmitters involved, specific receptor blockers of NMDA (AP-5), GABAA (bicuculline, Bic) and Glycine (strychnine, Str) were applied locally. Results GON stimulation produced a facilitation of the response to light facial mechanical stimuli in controls, and an inhibition in CCI-IoN cases. AP-5 reduced responses to GON and vibrissal stimulation and blocked the facilitation of GON on vibrissal responses found in controls. The application of Bic or Str significantly reduced the facilitatory effect of GON stimulation on the response to vibrissal stimulation in controls. However, the opposite effect was found when GABAergic or Glycinergic transmission was prevented in CCI-IoN cases. Conclusions GON stimulation modulates the responses of TCC neurons to light mechanical input from the face in opposite directions in controls and under CCI-IoN. This modulation is mediated by GABAergic and Glycinergic mechanisms. These results will help to elucidate the neural mechanisms underlying the effectiveness of nerve stimulation in controlling painful craniofacial disorders, and may be instrumental in identifying new therapeutic targets for their prevention and treatment.post-print1932 K

Functional Electrical Stimulation of Peripheral Nerve Tissue Via Regenerative Sieve Microelectrodes

Functional electrical stimulation (FES) of peripheral nervous tissue offers a promising method for restoring motor function in patients suffering from complex neurological injuries. However, existing microelectrodes designed to stimulate peripheral nerve are unable to provide the type of stable, selective interface required to achieve near-physiologic control of peripheral motor axons and distal musculature. Regenerative sieve electrodes offer a unique alternative to such devices, achieving a highly stable, selective electrical interface with independent groups of regenerated nerve fibers integrated into the electrode. Yet, the capability of sieve electrodes to functionally recruit regenerated motor axons for the purpose of muscle activation remains largely unexplored. The present dissertation aims to examine the potential role of regenerative electrodes in FES applications by testing the unifying hypothesis that sieve electrodes of various design and geometry are capable of selectively stimulating regenerated motor axons for the purpose of controlling muscle activation. This hypothesis was systematically tested through a series of experiments examining the ability of both micro-sieve electrodes and macro-sieve electrodes to achieve a stable interface with peripheral nerve tissue, electrically activate small groups of regenerated motor axons, and selectively recruit motor units present in multiple distal muscles. Custom sieve electrodes were fabricated via sacrificial photolithography. In vivo testing in rat sciatic nerve validated the ability of chronically-implanted regenerative sieve electrodes to support motor axon regeneration and integrate into peripheral nerve tissue. Sieve electrode geometry was shown to strongly modulate axonal regeneration, muscle reinnervation, and device functionality, as high-transparency macro-sieve electrodes facilitated superior neural integration and functional recovery compared to low-transparency micro-sieve electrodes. Inclusion of neurotrophic factors into sieve electrode assemblies increased axonal regeneration through implanted electrodes and improved the quality of the sieve/nerve interface in low-transparency devices. In vivo testing in rat sciatic nerve further validated the ability of chronically-implanted regenerative sieve electrodes to facilitate FES of regenerated motor axons and selective recruitment of distal musculature. Selective stimulation of regenerated motor axons using implanted micro- and macro-sieve electrodes enabled effective, external control of muscle activation within anterior and posterior compartments of the lower leg (e.g. ankle plantarflexion / dorsiflexion). Selective activation of distal musculature was achieved through modulation of stimulus amplitude, channel activation, and field steering. In summary, the present body of work provides initial evidence of the utility of regenerative electrodes as a means of selectively interfacing peripheral nerve tissue for the purpose of restoring muscle activation and motor control. These findings further highlight the clinical potential of implantable microelectrodes capable of intimately integrating into host neural tissue

Characterisation of interneurons in lamina II of the rat spinal cord

Lamina II of the dorsal horn contains numerous small neurons with varying morphologies, most of which have axons that remain within the spinal cord. It can be distinguished from the other laminae by its lack of myelinated fibres and its constituent interneurons that are densely packed. This region is the major termination site for unmyelinated (C) primary afferent fibres, which convey mostly nociceptive information. It also receives inputs from thinly myelinated (Aδ) fibres, some of which are nociceptive. In spite of its importance and several past attempts, little is known of its neuronal circuitry. This is mainly due to the great functional and morphological diversity of lamina II interneurons, which has made characterisation difficult. A comprehensive classification scheme is essential to identify discrete functional populations of lamina II interneurons, and to enable understanding of their roles in the local neuronal circuitry. The present study aims to investigate the physiological, pharmacological and morphological properties of lamina II interneurons recorded in an in vitro slice preparation from adult rat spinal cord. These properties were correlated with the neurotransmitter content of each cell, which was identified by detection of vesicular transporters in axonal boutons, in order to distinguish discrete functional subpopulations of cells in this region. Both inhibitory and excitatory interneurons were identified in lamina II, based on their expression of vesicular GABA transporter (VGAT) or vesicular glutamate transporter (VGLUT2), respectively. None of the cells that had VGAT-immunoreactive axons displayed staining for VGLUT2, and vice-versa. Injection of depolarising current evoked tonic-, transient-, delayed-, gap-, reluctant- and single spike-firing among these cells. Discharge pattern was strongly related to neurotransmitter phenotype, since most excitatory cells, but very few inhibitory cells had firing patterns that could be attributed to A-type potassium (IA) currents (i.e. delayed, gap or reluctant-firing). This suggests that excitatory lamina II interneurons with IA –type firing patterns are involved in plasticity that contributes to pain states. The majority of inhibitory cells displayed tonic-firing pattern in response to depolarisation. There was also an obvious difference in the response of lamina II neurons to hyperpolarisation, since the majority of inhibitory cells showed inward currents while most excitatory cells displayed transient outward currents. Noradrenaline and serotonin hyperpolarised both inhibitory and excitatory neurons, while only inhibitory neurons responded to somatostatin. This is consistent with the findings of a previous study that had shown that the somatostatin 2 receptor (sst2a) is only expressed by inhibitory neurons in lamina II, and suggests that the pro-nociceptive effects of somatostatin are mediated by ‘disinhibition’. The somatodendritic morphology of 61 lamina II interneurons was reconstructed from projected confocal images of Neurobiotin labelling and assessed according to the morphological scheme developed by Grudt and Perl (2002). Although cells in the islet, central, vertical and radial class were identified, a substantial number of cells (19/61) had morphology that was atypical or intermediate between two classes and therefore could not be classified. Certain morphological types were consistently found in the inhibitory or excitatory population: all islet cells were GABAergic, while all radial cells and most vertical cells were glutamatergic. However, the correlation between these properties may be complex, since there was a considerable diversity in the remaining cells. Some glutamatergic interneurons had axons that contained somatostatin and many of these also contained enkephalin. Somatostatin-expressing glutamatergic cells included various morphological types, while enkephalin was detected in the axons of vertical and radial cells. All cells with axons that were somatostatin- and enkephalin-immunoreactive had delayed-firing patterns. Taken together with the pharmacological data from the present study, this suggests that somatostatin released from these glutamatergic neurons would hyperpolarise subsets of inhibitory neurons and causes disinhibition. This could lead to alterations of pain thresholds. The results from this study demonstrate that distinctive populations of inhibitory and excitatory interneurons can be recognised in lamina II, and these cells are most likely to correspond to discrete functional groups. Electrophysiological, neurochemical, morphological and pharmacological properties of neurons can be correlated but this is likely to be very complex. Future investigations that combine various approaches should allow further understanding of the specific roles of lamina II interneurons in nociceptive processing within the spinal cord

Axonal Ensheathment and Intercellular Barrier Formation in Drosophila

Glial cells are critical players in every major aspect of nervous system development, function, and disease. Other than their traditional supportive role, glial cells perform a variety of important functions such as myelination, synapse formation and plasticity, and establishment of blood–brain and blood–nerve barriers in the nervous system. Recent studies highlight the striking functional similarities between Drosophila and vertebrate glia. In both systems, glial cells play an essential role in neural ensheathment thereby isolating the nervous system and help to create a local ionic microenvironment for conduction of nerve impulses. Here, we review the anatomical aspects and the molecular players that underlie ensheathment during different stages of nervous system development in Drosophila and how these processes lead to the organization of neuroglial junctions. We also discuss some key aspects of the invertebrate axonal ensheathment and junctional organization with that of vertebrate myelination and axon–glial interactions. Finally, we highlight the importance of intercellular junctions in barrier formation in various cellular contexts in Drosophila. We speculate that unraveling the genetic and molecular mechanisms of ensheathment across species might provide key insights into human myelin-related disorders and help in designing therapeutic interventions