Sensory hypersensitivity occurs soon after whiplash injury and is associated with poor recovery

Hypersensitivity to a variety of sensory Stimuli is a feature of persistent whiplash associated disorders (WAD). However, little is known about sensory disturbances from the time Of injury until transition to either recovery or symptom persistence. Quantitative sensory testing (pressure and thermal pain thresholds, the brachial plexus provocation test), the sympathetic vasoconstrictor reflex and psychological distress (GHQ-28) were prospectively measured in 76 whiplash Subjects within 1 month of injury and then 2, 3 and 6 months post-injury. Subjects were classified at 6 months post-injury using scores on the Neck Disability Index: recovered (30). Sensory and sympathetic nervous system tests were also measured in 20 control subjects. All whiplash groups demonstrated local mechanical hyperalgesia in the cervica spine at 1 month post-injury. This hyperalgesia persisted in those with moderate/severe symptoms at 6 months but resolved by 2 months in those who had recovered or reported persistent mild symptoms. Only those with persistent moderate/severe symptoms at 6 months demonstrated generalised hypersensitivity to all sensory tests. These changes Occurred within 1 month of injury and remained Unchanged throughout the Study period. Whilst no significant group differences were evident for the sympathetic vasoconstrictor response, the moderate/severe group showed a tendency for diminished sympathetic reactivity. GHQ-28 scores of the moderate/severe group were higher than those of the other two groups. The differences in GHQ-28 did not impact on any of the sensory measures. These findings suggest that those with persistent moderate/severe symptoms at 6 months display, soon after injury, generalised hypersensitivity suggestive of changes in central pain processing mechanisms. This phenomenon did not Occur in those who recover or those with persistent mild symptoms. (C) 2003 International Association for the Study of Pain. Published by Elsevier Science B.V. All rights reserved.

Characterization of acute whiplash-associated disorders

Study Design. An experimental study of motor and sensory function and psychological distress in subjects with acute whiplash injury. Objectives. To characterize acute whiplash injury in terms of motor and sensory systems dysfunction and psychological distress and to compare subjects with higher and lesser levels of pain and disability. Summary of Background Data. Motor system dysfunction, sensory hypersensitivity, and psychological distress are present in chronic whiplash associated disorders ( WAD), but little is known of such factors in the acute stage of injury. As higher levels of pain and disability in acute WAD are accepted as signs of poor outcome, further characterization of this group from those with lesser symptoms is important. Materials and Methods. Motor function ( cervical range of movement [ ROM], joint position error [JPE]; activity of the superficial neck flexors [EMG] during a test of craniocervical flexion), quantitative sensory testing ( pressure, thermal pain thresholds, and responses to the brachial plexus provocation test), and psychological distress (GHQ-28, TAMPA, IES) were measured in 80 whiplash subjects ( WAD II or III) within 1 month of injury, as were 20 control subjects. Results. Three subgroups were identified in the cohort using cluster analysis based on the Neck Disability Index: those with mild, moderate, or severe pain and disability. All whiplash groups demonstrated decreased ROM and increased EMG compared with the controls ( all P < 0.01). Only the moderate and severe groups demonstrated greater JPE and generalized hypersensitivity to all sensory tests ( all P < 0.01). The three whiplash subgroups demonstrated evidence of psychological distress, although this was greater in the moderate and severe groups. Measures of psychological distress did not impact on between group differences in motor or sensory tests. Conclusions. Acute whiplash subjects with higher levels of pain and disability were distinguished by sensory hypersensitivity to a variety of stimuli, suggestive of central nervous system sensitization occurring soon after injury. These responses occurred independently of psychological distress. These findings may be important for the differential diagnosis of acute whiplash injury and could be one reason why those with higher initial pain and disability demonstrate a poorer outcome.

Type 1 nitrergic (ND1) cells of the rabbit retina: Comparison with other axon-bearing amacrine cells

NADPH diaphorase (NADPHd) histochemistry labels two types of nitrergic amacrine cells in the rabbit retina. Both the large ND1 cells and the small ND2 cells stratify in the middle of the inner plexiform layer, and their overlapping processes produce a dense plexus, which makes it difficult to trace the morphology of single cells. The complete morphology of the ND1 amacrine cells has been revealed by injecting Neurobiotin into large round somata in the inner nuclear layer, which resulted in the labelling of amacrine cells whose proximal morphology and stratification matched those of the ND1 cells stained by NADPHd histochemistry. The Neurobiotin-injected ND1 cells showed strong homologous tracer coupling to surrounding ND1 cells, and double-labelling experiments confirmed that these coupled cells showed NADPHd reactivity. The ND1 amacrine cells branch in stratum 3 of the inner plexiform layer, where they produce a sparsely branched dendritic tree of 400-600 mum diameter in ventral peripheral retina. In addition, each cell gives rise to several fine beaded processes, which arise either from a side branch of the dendritic tree or from the tapering of a distal dendrite. These axon-like processes branch successively within the vicinity of the dendritic field before extending, with little or no further branching, for 3-5 mm from the soma in ventral peripheral retina. Consequently, these cells may span one-third of the visual field of each eye, and their spatial extent appears to be greater than that of most other types of axon-bearing amacrine cells injected with Neurobiotin in this study. The morphology and tracer-coupling pattern of the ND1 cells are compared with those of confirmed type 1 catecholaminergic cells, a presumptive type 2 catecholaminergic cell, the type 1 polyaxonal. cells, the long-range amacrine cells, a novel type of axon-bearing cell that also branches in stratum 3, and a type of displaced amacrine cell that may correspond to the type 2 polyaxonal cell. (C) 2004 Wiley-Liss, Inc.

The type 1 polyaxonal amacrine cells of the rabbit retina: A tracer-coupling study

The type 1 polyaxonal (PA1) cell is a distinct type of axon-bearing amacrine cell whose soma commonly occupies an interstitial position in the inner plexiform layer; the proximal branches of the sparse dendritic tree produce 1-4 axon-like processes, which form an extensive axonal arbor that is concentric with the smaller dendritic tree (Dacey, 1989; Famiglietti, 1992a,b). In this study, intracellular injections of Neurobiotin have revealed the complete dendritic and axonal morphology of the PA1 cells in the rabbit retina, as well as labeling the local array of PA1 cells through homologous tracer coupling. The dendritic-field area of the PA1 cells increased from a minimum of 0.15 mm(2) (0.44-mm equivalent diameter) on the visual streak to a maximum of 0.67 mm(2) (0.92-mm diameter) in the far periphery; the axonal-field area also showed a 3-fold variation across the retina, ranging from 3.1 mm(2) (2.0-mm diameter) to 10.2 mm(2) (3.6-mm diameter). The increase in dendritic- and axonal-field size was accompanied by a reduction in cell density, from 60 cells/mm(2) in the visual streak to 20 cells/mm(2) in the far periphery, so that the PA1 cells showed a 12 times overlap of their dendritic fields across the retina and a 200-300 times overlap of their axonal fields. Consequently, the axonal plexus was much denser than the dendritic plexus, with each square millimeter of retina containing similar to100 mm of dendrites and similar to1000 mm of axonal processes. The strong homologous tracer coupling revealed that similar to45% of the PA1 somata were located in the inner nuclear layer, similar to50% in the inner plexiform layer, and similar to5% in the ganglion cell layer. In addition, the Neurobiotin-injected PA1 cells sometimes showed clear heterologous tracer coupling to a regular array of small ganglion cells, which were present at half the density of the PA1 cells. The PA1 cells were also shown to contain elevated levels of gamma-aminobutyric acid (GABA), like other axon-bearing amacrine cells.

Vasodilator mechanisms in the dorsal aorta of the giant shovelnose ray, Rhinobatus typus (Rajiformes; Rhinobatidae)

This study investigated the nature of vasodilator mechanisms in the dorsal aorta of the giant shovelnose ray, Rhinobatus typus. Anatomical techniques found no evidence for an endothelial nitric oxide synthase, but neural nitric oxide synthase was found to be present in the perivascular nerve fibres of the dorsal aorta and other arteries and veins using both NADPH-diaphorase staining and immunohistochemistry with a specific neural NOS antibody. Arteries and veins both contained large nNOS-positive nerve trunks from which smaller nNOS-positive bundles branched and formed a plexus in the vessel wall. Single, varicose nNOS-positive nerve fibres were present in both arteries and veins. Within the large bundles of both arteries and veins, groups of nNOS-positive cell bodies forming microganglia were observed. Double-labelling immunohistochemistry using an antibody to tyrosine hydroxylase showed that nearly all the NOS nerves were not sympathetic. Acetylcholine always caused constriction of isolated rings of the dorsal aorta and the nitric oxide donor, sodium nitroprusside, did not mediate any dilation. Addition of nicotine (3 x 10(-4) M) to preconstricted rings caused a vasodilation that was not affected by the nitric oxide synthase inhibitor, L-NNA (10(-4) M), nor the soluble guanylyl cyclase inhibitor, ODQ (10(-5) M). This nicotine-mediated vasodilation was, therefore, not due to the synthesis and release of NO. Disruption of the endothelium significantly reduced or eliminated the nicotine-mediated vasodilation. In addition. indomethacin (10(-5) M), an inhibitor of cyclooxygenases, significantly increased the time period to maximal dilation and reduced, but did not completely inhibit the nicotine-mediated vasodilation. These data support the hypothesis that a prostaglandin is released from the vascular endothelium of a batoid ray, as has been described previously in other groups of fishes. The function of the nitrergic innervation of the blood vessels is not known because nitric oxide does not appear to regulate vascular tone. (C) 2003 Elsevier Inc. All rights reserved.

Developmental changes in expression of GABA(A) receptor-channels in rat intrinsic cardiac ganglion neurones

The effects of gamma-aminobutyric acid (GABA) on the electrophysiological properties of intracardiac neurones were investigated in the intracardiac ganglion plexus in situ and in dissociated neurones from neonatal, juvenile and adult rat hearts. Focal application of GABA evoked a depolarizing, excitatory response in both intact and dissociated intracardiac ganglion neurones. Under voltage clamp, both GABA and muscimol elicited inward currents at -60 mV in a concentration-dependent manner. The fast, desensitizing currents were mimicked by the GABA(A) receptor agonists muscimol and taurine, and inhibited by the GABA(A) receptor antagonists, bicuculline and picrotoxin. The GABA(A0) antagonist (1,2,5,6-tetrahydropyridin-4-yl)methyl phosphonic acid (TPMPA), had no effect on GABA-induced currents, suggesting that GABA(A) receptor-channels mediate the response. The GABA-evoked current amplitude recorded from dissociated neurones was age dependent whereby the peak current density measured at -100 mV was similar to 20 times higher for intracardiac neurones obtained from neonatal rats (P2-5) compared with adult rats (P45-49). The decrease in GABA sensitivity occurred during the first two postnatal weeks and coincides with maturation of the sympathetic innervation of the rat heart. Immunohistochemical staining using antibodies against GABA demonstrate the presence of GABA in the intracardiac ganglion plexus of the neonatal rat heart. Taken together, these results suggest that GABA and taurine may act as modulators of neurotransmission and cardiac function in the developing mammalian intrinsic cardiac nervous system.

Physical and psychological factors predict outcome following whiplash injury

Predictors of outcome following whiplash injury are limited to socio-demographic and symptomatic factors, which are not readily amenable to secondary and tertiary intervention. This prospective study investigated the predictive capacity of early measures of physical and psychological impairment on pain and disability 6 months following whiplash injury. Motor function (ROM; kinaesthetic sense; activity of the superficial neck flexors (EMG) during cranio-cervical flexion), quantitative sensory testing (pressure, thermal pain thresholds, brachial plexus provocation test), sympathetic vasoconstrictor responses and psychological distress (GHQ-28, TSK, IES) were measured in 76 acute whiplash participants. The outcome measure was Neck Disability Index scores at 6 months. Stepwise regression analysis was used to predict the final NDI score. Logistic regression analyses predicted membership to one of the three groups based on final NDI scores (< 8 recovered, 10-28 mild pain and disability, > 30 moderate/severe pain and disability). Higher initial NDI score (1.007-1.12), older age (1.03-1.23), cold hyperalgesia (1.05-1.58), and acute post-traumatic stress (1.03-1.2) predicted membership to the moderate/severe group. Additional variables associated with higher NDI scores at 6 months on stepwise regression analysis were: ROM loss and diminished sympathetic reactivity. Higher initial NDI score (1.03-1.28), greater psychological distress (GHQ-28) (1.04-1.28) and decreased ROM (1.03-1.25) predicted subjects with persistent milder symptoms from those who fully recovered. These results demonstrate that both physical and psychological factors play a role in recovery or non-recovery from whiplash injury. This may assist in the development of more relevant treatment methods for acute whiplash. (c) 2004 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

Strain and excursion of the sciatic, tibial, and plantar nerves during a modified straight leg raising test

A modified straight leg raising (SLR) in which ankle dorsiflexion is performed before hip flexion has been suggested to diagnose distal neuropathies such as tarsal tunnel syndrome. This study evaluates the clinical hypothesis that strain in the nerves around the ankle and foot caused by ankle dorsiflexion can be further increased with hip flexion. Linear displacement transducers were inserted into the sciatic, tibial, and plantar nerves and plantar fascia of eight embalmed cadavers to measure strain during the modified SLR. Nerve excursion was measured with a digital calliper. Ankle dorsiflexion resulted in a significant strain and distal. excursion of the tibial nerve. With the ankle in dorsiflexion, the proximal excursion and tension increase in the sciatic nerve associated with hip flexion were transmitted distally along the nerve from the hip to beyond the ankle. As hip flexion had an impact on the nerves around the ankle and foot but not on the plantar fascia, the modified SLR may be a useful test to differentially diagnose plantar heel pain. Although the modified SLR caused the greatest increase in nerve strain nearest the moving joint, mechanical forces acting on peripheral nerves are transmitted well beyond the moving joint. (c) 2006 Orthopaedic Research Society.

Physical and psychological factors maintain long-term predictive capacity post-whiplash injury

Higher initial levels of pain and disability, older age, cold hyperalgesia, impaired sympathetic vasoconstriction and moderate post-traumatic stress symptoms have been shown to be associated with poor outcome 6 months following whiplash injury. This study prospectively investigated the predictive capacity of these variables at a long-term follow-up. Sixty-five of an initial cohort of 76 acutely injured whiplash participants were followed to 2-3 years post-accident. Motor function (ROM; kinaesthetic sense; activity of the superficial neck flexors (EMG) during cranio-cervical flexion), quantitative sensory testing (pressure, thermal pain thresholds and brachial plexus provocation test), sympathetic vasoconstrictor responses and psychological distress (GHQ-28, TSK and IES) were measured. The outcome measure was Neck Disability Index (NDI) scores. Participants with ongoing moderate/severe symptoms at 2-3 years continued to manifest decreased ROM, increased EMG during cranio-cervical flexion, sensory hypersensitivity and elevated levels of psychological distress when compared to recovered participants and those with milder symptoms. The latter two groups showed only persistent deficits in cervical muscle recruitment patterns. Higher initial NDI scores (OR 1.00-1.1), older age (OR 1.00-1.13), cold hyperalgesia (OR 1.1-1.13) and post-traumatic stress symptoms (OR 1.03-1.2) remained significant predictors of poor outcome at long-term follow-up (r(2) = 0.56). The robustness of these physical and psychological factors suggests that their assessment in the acute stage following whiplash injury will be important. (c) 2006 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

Implantation of a scaffold following bulbectomy induces laminar organization of regenerating olfactory axons

Primary olfactory axons expressing different odorant receptors are interspersed within the olfactory nerve. However, upon reaching the outer nerve fiber layer of the olfactory bulb they defasciculate, sort out, and refasciculate prior to targeting glomeruli in fixed topographic positions. While odorant receptors are crucial for the final targeting of axons to glomeruli, it is unclear what directs the formation of the nerve fiber and glomerular layers of the olfactory bulb. While the olfactory bulb itself may provide instructive cues for the development of these layers, it is also possible that the incoming axons may simply require the presence of a physical scaffold to establish the outer laminar cytoarchitecture. In order to begin to understand the underlying role of the olfactory bulb in development of the outer layers of the olfactory bulb, we physically ablated the olfactory bulbs in OMP-IRES-LacZ and P2-IRES-tau-LacZ neonatal mice and replaced them with artificial biological scaffolds molded into the shape of an olfactory bulb. Regenerating axons projected around the edge of the cranial cavity at the periphery of the artificial scaffold and were able to form an olfactory nerve fiber layer and, to some extent, a glomerular layer. Our results reveal that olfactory axons are able to form rudimentary cytoarchitectonic layers if they are provided with an appropriately shaped biological scaffold. Thus, the olfactory bulb does not appear to provide any tropic substance that either attracts regenerating olfactory axons into the cranial cavity or induces these axons to form a plexus around its outer surface. (c) 2006 Elsevier B.V. All rights reserved.

The sorting behaviour of olfactory and vomeronasal axons during regeneration

In order to begin to understand how primary olfactory and vomeronasal organ (VNO) axons target specific regions of the olfactory bulb, we examined the sorting behaviour of these axons following neonatal unilateral olfactory bulbectomy. Bulbectomy induced widespread ipsilateral death of the primary olfactory and VNO neurons. After 4 weeks, many new sensory axons had re-grown into the cranial cavity and established a prominent plexus with evidence of dense tufts that were similar in gross appearance to glomeruli. Axons expressing the cell adhesion molecule OCAM, which normally innervate the ventrolateral and rostral halves of the main and accessory olfactory bulbs, respectively, sorted out and segregated from those axons not expressing this molecule within the plexus. In addition, VNO axons formed large discrete bundles that segregated from main olfactory axons within the plexus. Thus, VNO and primary olfactory axons as well as discrete subpopulations of both are able to sort out and remain segregated in the absence of the olfactory bulb. Sorting and convergence of axons therefore occur independently of the olfactory bulb and are probably attributable either to inherent properties of the axons themselves or to interactions between the axons and accompanying glial ensheathing cells.

Comparison and characterisation of ribozyme delivery in cultured glial and neuronal cells in vivo and in vitro

Ribozymes are short strands of RNA that possess a huge potential as biological tools for studying gene expression and as therapeutic agents to down-regulate undesirable gene expression. Successful application of ribozymes requires delivery to the target site in sufficient amounts for an adequate duration. However, due to their large size and polyanionic character ribozymes are not amenable to transport across biological membranes. In this study a chemically modified ribozyme with enhanced biological stability, targeted against the EGFR mRNA has been evaluated for cellular delivery to cultured glial and neuronal cells with a view to developing treatments for brain tumours. Cellular delivery of free ribozyme was characterised in cultured glial and neuronal cells from the human and rat. Delivery was very limited and time dependent with no consistent difference observed between glial and neuronal cells in both species. Cellular association was largely temperature and energy-dependent with a small component of non-energy dependent association. Further studies showed that ribozyme cellular association was inhibited with self and cross competition with nucleic and non-nucleic acid polyanions indicating the presence of cell surface ribozyme-binding molecules. Trypsin washing experiments further implied that the ribozyme binding surface molecules were protein by nature. Dependence of cellular association on pH indicated that interaction of ribozyme with cell surface molecules was based on ionic interactions. Fluoresence studies indicated that, post cell association, ribozymes were sequestered in sub-cellular vesicles. South-Western blots identified several cell surface proteins which bind to ribozymes and could facilitate cellular association. The limited cellular association observed with free ribozyme required the development and evaluation of polylactide-co-glycolide microspheres incorporating ribozyme for enhanced cellular delivery. Characterisation of microsphere mediated delivery of ribozyme in cultured glial and neuronal cells showed that association increased by 18 to 27-fold in all cell types with no differences observed between cell lines and species. Microsphere mediated delivery was temperature and energy dependent and independent of pH. In order to assess the potential of PLGA micro spheres for the CNS delivery of ribozyme the distribution of ribozyme entrapping microspheres was investigated in rat CNS after intracerebroventricular injection. Distribution studies demonstrated that after 24 hours there was no free ribozyme present in the brain parenchyma, however microsphere entrapped ribozyme was found in the CNS. Microspheres remained in the ventricular system after deposition and passed from the lateral ventricles to the third and fourth ventricle and in the subarachnoid space. Investigation of the influence of microsphere size on the distribution in CNS demonstrated that particles up to 2.5 and O.5f.lm remained in the ventricles around the choroid plexus and ependymal lining.

Phytoestrogens modulate breast cancer resistance protein expression and function at the blood-cerebrospinal fluid barrier

PURPOSE: Breast cancer resistance protein (BCRP/ABCG2) is a drug efflux transporter expressed at the blood cerebrospinal fluid barrier (BCSFB), and influences distribution of drugs into the central nervous systems (CNS). Current inhibitors have failed clinically due to neurotoxicity. Novel approaches are needed to identify new modulators to enhance CNS delivery. This study examines 18 compounds (mainly phytoestrogens) as modulators of the expression/function of BCRP in an in vitro rat choroid plexus BCSFB model. METHODS: Modulators were initially subject to cytotoxicity (MTT) assessment to determine optimal non-toxic concentrations. Reverse-transcriptase PCR and confocal microscopy were used to identify the presence of BCRP in Z310 cells. Thereafter modulation of the intracellular accumulation of the fluorescent BCRP probe substrate Hoechst 33342 (H33342), changes in protein expression of BCRP (western blotting) and the functional activity of BCRP (membrane insert model) were assessed under modulator exposure. RESULTS: A 24 hour cytotoxicity assay (0.001 µM-1000 µM) demonstrated the majority of modulators possessed a cellular viability IC50 > 148 µM. Intracellular accumulation of H33342 was significantly increased in the presence of the known BCRP inhibitor Ko143 and, following a 24 hour pre-incubation, all modulators demonstrated statistically significant increases in H33342 accumulation (P < 0.001), when compared to control and Ko143. After a 24 hour pre-incubation with modulators alone, a 0.16-2.5-fold change in BCRP expression was observed for test compounds. The functional consequences of this were confirmed in a permeable insert model of the BCSFB which demonstrated that 17-β-estradiol, naringin and silymarin (down-regulators) and baicalin (up-regulator) can modulate BCRP-mediated transport function at the BCSFB. CONCLUSION: We have successfully confirmed the gene and protein expression of BCRP in Z310 cells and demonstrated the potential for phytoestrogen modulators to influence the functionality of BCRP at the BCSFB and thereby potentially allowing manipulation of CNS drug disposition.

Origem do plexo lombossacral e seus nervos em quatis (Nasua nasua, LINNAEUS, 1766)

The objective of this research is to study the origin of the lumbosacral plexus and its nerves in coatis (Nasua nasua). Six adult specimens, three males and three females, were used in this research. The animals were donated by IBAMA / GO or were collected when found dead by the edges of highways in Southeast Goiás. The specimens were fixed in aqueous 10% formaldehyde solution and preserved in the same solution. The observations reveal that the lumbosacral plexus of the coati has a structure with few intercommunications between its components. There are no typical handle formations, but the roots or its branches converge to form the nerves that are driven to the pelvic limb. The components of the Lumbosacral Plexus are ventral branches of L4 to S3. The Lumbosacral plexus generates several short nerves destined to pelvic structures and, at the same time, three long nerves: femoral, obturator and sciatic, all destined to the hind limb.