Neuronal degeneration associated with sympathosensory plexuses in the trigeminal ganglia of aged mice that overexpress nerve growth factor.

Aberrant sympathetic sprouting is seen in the uninjured trigeminal ganglia of transgenic mice that ectopically express nerve growth factor under the control of the glial fibrillary acidic protein promoter. These sympathetic axons form perineuronal plexuses around a subset of sensory somata in 2- to 3-month-old transgenic mice. Here, we show that aged transgenic mice (i.e., 11-14 and 16-18 months old) have dystrophic sympathetic plexuses (i.e., increased densities of swollen axons), and that satellite glial cells, specifically those in contact with dystrophic plexuses in the aged mice display strong immunostaining for tumor necrosis factor alpha. The colocalization of dystrophic plexuses and reactive satellite glial cells in the aged mice coincides with degenerative features in the enveloped sensory somata. Collectively, these novel results show that, with advancing age, sympathetic plexuses undergo dystrophic changes that heighten satellite glial cell reactivity and that together these cellular events coincide with neuronal degeneration.

The association of metal ion exposure with alpha-synuclein-like immunoreactivity in the central nervous system of fish, Catostomus commersoni.

Alpha-synuclein protein aggregates are a major component of Lewy bodies, the intracytoplasmic inclusions found in dopaminergic neurons that are a defining characteristic of Parkinson's disease. Other "synucleopathies" include dementia with Lewy bodies and multisystem atrophy. In vitro, the formation of these deposits can be induced by a number of substances, including metal ions. Fish provide a useful model to study the long-term biological effects of metal ion exposure, but to date no studies have been reported concerning such exposures with respect to alpha-synuclein aggregation. Mature white sucker fish (Catostomus commersoni; aged 5-8 years) were sampled from two sites within the Red Lake area of Northwestern Ontario, a region highly contaminated by metal ions due to mining activity. Individual fish were characterized with respect to liver metal ion uptake and metallothionein levels. Central nervous system (CNS) tissues of fish from test sites representing high and low metal ion contamination were examined immunohistochemically using a polyclonal antibody recognising alpha-synuclein protein. We demonstrate here that the CNS of fish exposed to elevated metal ion environments had increased alpha-synuclein-like immunoreactive aggregates, potentially reflecting metal ion exposure leading to CNS toxicity. These findings demonstrate that fish may be an important new model for studying environmental risk factors and the pathology associated with Parkinson's disease.

Conformational analysis of the effects of methylmercury on nerve growth factor and brain derived neurotrophic factor.

Methylmercury is a neurotoxicant that is detrimental to the development and physiology of the nervous system. One possible mechanism for methylmercury's toxicity stems from its ability to interfere with the signaling of the neurotrophins nerve growth factor and brain derived neurotrophic factor. In this study, we examine the effect of methylmercury to determine if it interferes with neurotrophin conformation in a manner similar to Hg(2+), or if it occurs via an alternate mechanism. Our findings indicate that although MeHg inhibits neurotrophin signaling, its toxic effects are not mediated via an induced conformational change, as seen with other metal ions, including Hg(2+).

Null mutations for exon III and exon IV of the p75 neurotrophin receptor gene enhance sympathetic sprouting in response to elevated levels of nerve growth factor in transgenic mice.

Under normal conditions, expression of the p75 neurotrophin receptor (p75NTR) by sympathetic neurons can increase the affinity of the signaling receptor, trkA, to target-derived nerve growth factor (NGF) at distal axons. We have previously reported that sprouting of sympathetic axons into NGF-rich target tissues is enhanced when p75NTR expression is perturbed, leading to the postulate that p75NTR may restrain sympathetic sprouting in response to elevated NGF levels. These observations were made using mice having a null mutation of the third p75NTR exon, a line that may express a hypomorphic form of this receptor. Since mice carrying a null mutation of the fourth p75NTR exon may not express a similar splice variant, we sought to determine whether these animals possess the same phenotype of enhanced sympathetic sprouting in response to elevated levels of NGF. Both lines of transgenic mice lacking p75NTR displayed similar degrees of sympathetic axonal sprouting into the cerebellum and trigeminal ganglia, two target tissues having elevated levels of NGF protein. Furthermore, the densities of sympathetic axons in both targets were significantly greater than those observed in age-matched NGF transgenic siblings expressing full-length p75NTR. Our new findings provide a comparative analysis of the phenotype in two independent mutations of the same neurotrophin receptor, revealing that p75NTR plays an important role in restricting sympathetic sprouting in response to higher NGF levels.

Proteomic evaluation reveals that olfactory ensheathing cells but not Schwann cells express calponin.

Human clinical trials have begun worldwide that use olfactory ensheathing cells (OECs) to ameliorate the functional deficits following spinal cord injury. These trials have been initiated largely because numerous studies have reported that OECs transform into Schwann Cell (SC)-like cells that myelinate axons and support new growth in adult rats with spinal injury. This phenomenon is remarkable because OECs do not myelinate olfactory axons in their native environment. Furthermore, these myelinating OECs are morphologically identical to SCs, which can invade the spinal cord after injury. One factor that has contributed to a possible confusion in the identification of these cells is the lack of phenotypic markers to distinguish unequivocally between OECs and SCs. Such markers are required to first assess the degree of SC contamination in OEC cultures before intraspinal implantation, and then to accurately identify grafted OECs and invading SCs in the injured spinal cord. Using two-dimensional gel electrophoresis, we have identified calponin, an actin binding protein, as the first definitive phenotypic marker that distinguishes between OECs and SCs in vitro and in vivo. We have also provided ultrastructural evidence that calponin-immunopositive OECs do not transform into myelinating SC-like cells after intraspinal implantation. Rather, the grafted OECs retain their morphological and neurochemical features. These data yield new insight into the phenotypic characteristics of OECs, which together with invading SCs can enhance regeneration of the injured spinal cord.

Comparison of target innervation by sympathetic axons in adult wild type and heterozygous mice for nerve growth factor or its receptor trkA.

Nerve growth factor (NGF), a neurotrophin required for the survival and maintenance of postganglionic sympathetic neurons, mediates its trophic effects by activation of its high-affinity receptor trkA. Null mutant mice lacking either NGF or trkA have profound sympathetic deficits, thus revealing the vital importance of NGF synthesis in target tissues and trkA expression by sympathetic neurons. In this study, we sought to assess whether sympathetic neurons of the superior cervical ganglion (SCG) display alterations in their neurochemical phenotype in adult mice carrying one mutated allele for either NGF or trkA, and whether such differences result in altered patterns of innervation to the submandibular salivary gland and pineal gland. In comparison with adult siblings, levels of trkA protein in the SCG were reduced in age-matched NGF(+/-) and trkA(+/-) mice. While NGF(+/-) mice also had significantly fewer sympathetic axons innervating both the submandibular salivary gland and pineal gland, densities of sympathetic axons in both tissues reached normal levels in trkA(+/-) mice. These findings reveal that while levels of trkA are reduced in SCG neurons of adult NGF(+/-) and trkA(+/-) mice (compared with their wild type counterparts), sympathetic axons are capable of achieving normal patterns of target innervation in trkA(+/-) mice but not in NGF(+/-) mice. As NGF protein levels are not depleted in the submandibular salivary gland and pineal gland of NGF(+/-) mice, a loss of sympathetic neurons [Nat Neurosci 1999; 2:699-705], in combination with reduced levels of trkA protein, may account for perturbed patterns of sympathetic innervation to peripheral tissues.

Two TTX-resistant Na+ currents in mouse colonic dorsal root ganglia neurons and their role in colitis-induced hyperexcitability.

The composition of Na+ currents in dorsal root ganglia (DRG) neurons depends on their neuronal phenotype and innervation target. Two TTX-resistant (TTX-R) Na+ currents [voltage-gated Na channels (Nav)] have been described in small DRG neurons; one with slow inactivation kinetics (Nav1.8) and the other with persistent kinetics (Nav1.9), and their modulation has been implicated in inflammatory pain. This has not been studied in neurons projecting to the colon. This study examined the relative importance of these currents in inflammation-induced changes in a mouse model of inflammatory bowel disease. Colonic sensory neurons were retrogradely labeled, and colitis was induced by instillation of trinitrobenzenesulfonic acid (TNBS) into the lumen of the distal colon. Seven to ten days later, immunohistochemical properties were characterized in controls, and whole cell recordings were obtained from small (<40 pF) labeled DRG neurons from control and TNBS animals. Most neurons exhibited both fast TTX-sensitive (TTX-S)- and slow TTX-R-inactivating Na+ currents, but persistent TTX-R currents were uncommon (<15%). Most labeled neurons were CGRP (79%), tyrosine kinase A (trkA) (84%) immunoreactive, but only a small minority bind IB4 (14%). TNBS-colitis caused ulceration, thickening of the colon and significantly increased neuronal excitability. The slow TTX-R-inactivating Na current density (Nav1.8) was significantly increased, but other Na currents were unaffected. Most small mouse colonic sensory neurons are CGRP, trkA immunoreactive, but not isolectin B4 reactive and exhibit fast TTX-S, slow TTX-R, but not persistent TTX-R Na+ currents. Colitis-induced hyperexcitability is associated with increased slow TTX-R (Nav1.8) Na+ current. Together, these findings suggest that colitis alters trkA-positive neurons to preferentially increase slow TTX-R Na+ (Nav1.8) currents.

Structural and neurochemical features of postganglionic sympathetic neurons in the superior mesenteric ganglion of spontaneously hypertensive rats.

Postganglionic sympathetic neurons, which are exquisitely sensitive to small changes in levels of target-derived nerve growth factor (NGF), express two transmembrane receptors: 1) the trkA receptor mediates neuron survival and neurite outgrowth; and 2) the p75 neurotrophin receptor (p75NTR) enhances neuronal responsiveness of trkA to NGF. Elevating levels of NGF induces several morphological and neurochemical alterations in sympathetic neurons, including axonal sprouting, increased levels of p75NTR mRNA relative to trkA mRNA, and increased accumulations of NGF in hypertrophied somata. Spontaneously hypertensive rats (SHR) display both elevated NGF levels and increased sympathetic axonal innervation of the mesenteric vasculature. In this investigation we assessed whether sympathetic neurons innervating the mesenteric vasculature of SHR display other features indicative of increased levels of target-derived NGF. In 5-week-old SHR, levels of both p75NTR and trkA mRNA in mesenteric sympathetic neurons were significantly elevated compared to levels in age-matched control rats. By 15 and 30 weeks of age, levels of p75NTR mRNA expression in mesenteric sympathetic neurons were similar between SHR and control rats. Accumulations of NGF were depleted in the sympathetic somata of 15- and 30-week-old SHR compared to age-matched control rats. Moreover, sympathetic neurons in SHR were not hypertrophied, as the sizes of somata were comparable between SHR and control rats. Our data illustrate that despite having augmented levels of NGF in the mesenteric vasculature, SHR do not display many of the morphological and neurochemical features that are associated with an enhanced responsiveness by sympathetic neurons to elevated levels of target-derived NGF.