Characterization of a thy1.2 GFP transgenic mouse reveals a tissue-specific organization of the spinal dorsal horn.
ABSTRACT In this study, transgenic mice in which membrane-linked enhanced green fluorescent protein (mGFP) is expressed from the Thy1.2 promoter were used. In these mice, a subpopulation of small to medium sized DRG neurons double stained for IB4 but not for CGRP. Most of the peripheral terminals traversed the dermis and ramify within the epidermis and form superficial terminals. Within the spinal cord, these afferents terminated exclusively within the substantia gelatinosa (SG). A second fibre type in the skin also expressed mGFP, and formed club-shaped endings towards the bases of hairs. Injury to the sciatic nerve resulted in mGFP loss from the SG ipsilateral to the nerve injury, but also in the corresponding region contralaterally. Together, these findings reveal the specificity of connectivity of a defined subpopulation of DRG sensory neurons innervating the epidermis and this will facilitate analysis of their physiological functions.
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ABSTRACT: Two-photon microscopy enables high-resolution in vivo imaging of cellular morphology and activity, in particular of population activity in complex neuronal circuits. While two-photon imaging has been extensively used in a variety of brain regions in different species, in vivo application to the vertebrate spinal cord has lagged behind and only recently became feasible by adapting and refining the experimental preparations. A major experimental challenge for spinal cord imaging is adequate control of tissue movement, which meanwhile can be achieved by various means. One set of studies monitored structural dynamics of neuronal and glial cellular components in living animals using transgenic mice with specific expression of fluorescent proteins. Other studies employed in vivo calcium imaging for functional measurements of sensory-evoked responses in individual neurons of the dorsal horn circuitry, which at present is the only part of rodent spinal cord grey matter accessible for in vivo imaging. In a parallel approach, several research groups have applied two-photon imaging to sensorimotor circuits in the isolated spinal cord (in vitro) to provide complementary information and valuable new perspectives on the function of specific interneuron types in locomotor-related networks. In this review we summarize recent results from these types of high-resolution two-photon imaging studies in the spinal cord and provide experimental perspectives for improving and extending this approach in future applications.Experimental Neurology 07/2012; · 4.65 Impact Factor
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ABSTRACT: The skin is innervated by two populations of unmyelinated sensory fibers, the peptidergic and nonpeptidergic, which transmit nociceptive information to the central nervous system. The peptidergic population expresses neuropeptides such as substance P (SP) and calcitonin gene-related peptide (CGRP) and has both cutaneous and visceral targets. The nonpeptidergic population expresses the purinergic receptor P2X(3), binds the isolectin B4 (IB4), and innervates mainly the epidermis. To date, the peptidergic nociceptor population in cutaneous tissue of the rat has been well characterized, whereas the nonpeptidergic innervation pattern has lacked an adequate description. To this aim, we used light microscopic immunocytochemistry to investigate the pattern of P2X(3)-immunoreactive (-IR) fiber innervation of both hairy and glabrous skin from male Sprague-Dawley rats. Our results show extensive P2X(3)-IR fibers throughout the upper and lower dermis. Thick bundles of P2X(3)-IR fibers were found to run in parallel with the dermal-epidermal junction and projected multiple thin collateral axons that penetrated the epidermal layer, creating a dense network of innervation throughout the entire epidermis. The distribution of P2X(3)-IR fibers in the epidermis was far more extensive than the distribution of CGRP-IR fibers. P2X(3)-IR fibers also innervate hair follicles but were rarely found in close proximity to glands and blood vessels. The present results suggest a primary role for P2X(3)-IR fibers in the detection of noxious stimuli in cutaneous tissue and provide an anatomical basis for future studies examining a possible functionally distinct role of nonpeptidergic nociceptors in the transmission of nociceptive signals.The Journal of Comparative Neurology 04/2009; 514(6):555-66. · 3.66 Impact Factor
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ABSTRACT: Nitric oxide (NO) plays an important role in pathophysiology of the nervous system. Copper/zinc superoxide dismutase (SOD1) reacts with superoxide, which is also a substrate for NO, to provide antioxidative protection. NO production is greatly altered following nerve injury, therefore we hypothesised that SOD1 and NO may be involved in modulating axotomy responses in dorsal root ganglion (DRG)-spinal network. To investigate this interaction, adult Thy1.2 enhanced membrane-bound green fluorescent protein (eGFP) mice underwent sciatic nerve axotomy and received NG-nitro- <l-arginine methylester (L-NAME) or vehicle 7-9 days later. L4-L6 spinal cord and DRG were harvested for immunohistochemical analyses. Effect of injury was confirmed by axotomy markers; small proline-rich repeat protein 1A (SPRR1A) was restricted to ipsilateral neuropathology, while Thy1.2 eGFP revealed also contralateral crossover effects. L-NAME, but not axotomy, increased neuronal NO synthase (nNOS) and SOD1 immunoreactive neurons, with no colocalisation, in a lamina-dependent manner in the dorsal horn of the spinal cord. Axotomy and/or L-NAME had no effect on total nNOS+ and SOD1+ neurons in DRG. However, L-NAME altered SOD1 expression in subsets of axotomised DRG neurons. These findings provide evidence for differential distribution of SOD1 and its modulation by NO, which may interact to regulate axotomy-induced changes in DRG-spinal network.Neuron Glia Biology 05/2012; · 1.34 Impact Factor