Ronald Wiley - Academia.edu (original) (raw)

Papers by Ronald Wiley

Journal of Neuroscience Research, Jan 15, 1996

ABSTRACT Nerve growth factor (NGF) provides critical trophic support to the cholinergic basal for... more ABSTRACT Nerve growth factor (NGF) provides critical trophic support to the cholinergic basal forebrain neurons that express high levels of the low-affinity NGF receptor (p75NGFR) in the adult rat brain. Intraventricular injection of 192 IgG-saporin, made by coupling the monoclonal antibody to p75NGFR 192 IgG to the cytotoxin saporin, selectively destroys the p75NGFR-bearing neurons in the basal forebrain and was used here to examine the effects of selective cholinergic lesions on brain NGF protein levels. We showed that 192 IgG-saporin produced significant long-lasting elevation of NGF protein levels in the hippocampus, cortex, and olfactory bulb, with profound reductions of ChAT activities representing complete cholinergic deafferentations of these areas. NGF level was maintained in the basal forebrain, even though there was almost complete loss of p75NGFR-immunoreactive cells and significant decrease of ChAT activity. In addition, a mild glial response was observed in the basal forebrain, and most of the activated astroglia expressed NGF-like immunoreactivity there. The increases in NGF protein levels in the target areas of the basal forebrain were most likely due to loss of cholinergic basal forebrain neurons and retrograde transport of NGF from these areas. Glial-derived NGF is partially responsible for the maintained level of NGF in the basal forebrain after the loss of cholinergic neurons. The accumulation of NGF protein in the target areas may have some effects on synaptic rearrangement in denervated tissues.

J Neurosci Res, 1995

To characterize the specificity of a novel cholinergic immunotoxin (conjugate of the monoclonal a... more To characterize the specificity of a novel cholinergic immunotoxin (conjugate of the monoclonal antibody 192IgG against the low-affinity nerve growth factor receptor with the cytotoxic protein saporin), coronal sections through the basal forebrain of adult rats, that received a single intracerebro-ventricular injection of 4 micrograms of 192IgG-saporin conjugate, were subjected to histochemical and immunocytochemical procedures to evaluate cholinergic (choline acetyltransferase (ChAT)-immunoreactive, acetylcholinesterase-positive, NADPH-diaphorase-positive) and GABAergic structures (parvalbumin-immunoreactive, labeling of perineuronal nets with Wisteria floribunda agglutinin) as well as microglia (visualized with Griffonia simplicifolia agglutinin) and astrocytes (immunostaining for glial fibrillary acidic protein). Seven days following injection of the immunotoxin, ChAT-immunoreactive cells nearly completely disappeared throughout the magnocellular basal forebrain complex, including globus pallidus, as compared to vehicle-injected controls. However, there was no significant difference in the number of ChAT-positive cells in the adjacent ventral pallidum and in the caudate-putamen of immunolesioned and control animals. NADPH-diaphorase-containing cells, including a significant subpopulation of cholinergic cells, also strikingly decreased in number by more than 90% in the magnocellular basal forebrain complex following immunolesion, and only a few noncholinergic diaphorase-positive cells survived in the medial septum, vertical and horizontal diagonal band, and nucleus basalis of Meynert. In contrast, the number of parvalbumin-containing GABAergic projection neurons in the septum-diagonal band of Broca complex and nucleus basalis of Meynert from immunolesioned rats was not different from that of vehicle-injected control animals. Immunolesioning also did not result in any change in either number or shape of cells surrounded by perineuronal nets, which are frequently associated with parvalbumin-containing GABAergic neurons. Seven days following injection of the immunotoxin, a very strong activation of microglia with an identical distribution pattern was observed in all experimental animals. Large numbers of activated microglia were found in all magnocellular basal forebrain nuclei, corresponding to the distribution of degenerating cholinergic cells. Additionally, immunolesioning also resulted in a dramatic activation of microglia in the lateral septal nuclei, which are known to be almost free of cholinergic cells, but not of penetrating cholinergic dendrites in adjacent zones, and in the ventral pallidum, where there was no observed loss of cholinergic cells. There was no significant increase in microglia activation in striatum and cortical areas, and no astrocytic response in any of the basal forebrain nuclei at this particular time point of survival. These results suggest that 192IgG-saporin specifically destroys basal forebrain cholinergic neurons and does not suppress their neuronal activity.(ABSTRACT TRUNCATED AT 400 WORDS)

Brain Research, Jun 12, 1999

Removal of cholinergic septal inputs using the immunotoxin 192 IgG-saporin reduces the number of ... more Removal of cholinergic septal inputs using the immunotoxin 192 IgG-saporin reduces the number of interneurons containing Ž. w Ž. neuropeptide Y NPY-immunoreactivity in the rat dentate gyrus by approximately 30% Milner et al., J. Comp. Neurol. 386 1997 x 48-59. The goal of the present study was to determine if NPY-containing neurons that survive deafferentation have any distinguishing morphological andror microenvironmental features. For this, 2 or 24 weeks after intracerebroventricular injections of 192 IgG-saporin, NPY-immunolabeled neurons in the hilus of the dentate gyrus were examined by electron microscopy. Neither the size nor morphological traits of NPY-labeled perikaryal or dendritic profiles from lesioned compared to control rats at either time-point differed significantly. However, at both time-points, NPY-containing somatal profiles from immunolesioned rats compared to controls had a reduced percentage of their plasmalemmal surface apposed to unmyelinated axon profiles and an increased percentage of their surface occupied by astrocytic profiles. At the 24 week time-point, these differences were statistically significant. The primary contributing factor for these changes was Ž. the absence of a subgroup of NPY-labeled somatal profiles in lesioned rats compared to controls which was: a distinguished by frequent Ž. Ž. appositions of unmyelinated axons from 15 to 35% to the plasmalemmal surface; and b located primarily in the central hilar region. Unlike NPY-containing somata, changes associated with NPY-labeled dendritic profiles were exclusively related to associated presynaptic profiles at the 24 week time-point. In lesioned rats compared to controls at this time-point, NPY-containing dendritic profiles had a concurrent increase in the percentage of the plasmalemmal surface occupied by active zones and the size of terminals contacting them. Ž. The present results combined with those of our earlier study suggest that septal cholinergic deafferentation results in: a the loss of a Ž. distinct subpopulation of hippocampal NPY-containing neurons; and b an increase in total active zone area suggesting a strengthening of synaptic connections to the surviving population of NPY-containing neurons in the long term.

Brain Research, Dec 1, 1995

In the basal ganglia, centrally active suicide transport agents produce selective lesions of the ... more In the basal ganglia, centrally active suicide transport agents produce selective lesions of the striatopallidal and striatonigral pathways based on receptor binding and neuropeptide mRNA studies. Anatomical analyses indicate a selective, albeit modest, loss of projection neurons, in the present study, we sought to determine the ultrastructural sequelae in the stria/urn of suicide transport injections of the globus pallidus (GP) or substantia nigra (SN). Neostriata of adult rats were examined l(/ days after lesions of the striatopallidal or striatonigral pathways with OX7-saporin or volkensin. Controls consisted of normal unoperated rats and animals injected into either target with ricin, a toxic lectin that is not transported in the central nervous system. Injections with OX7-saporin or volkensin into the GP or SN produced a decrease in striatal synaptic density of approximately 20%, relative to the contralateral side. Dark degenerating profiles, though very rare in the contralateral striata, were present throughout the neuropil in the ipsilateral striata. In animals with striatopallidal lesions, axospinous synapses of both the asymmetric and symmetric type were decreased in density, while the number of synapses fl)rmed with dendritic shafts was unaffected. In addition, the number of striatal mitochondrial profiles was decreased ipsilateral to the lesions. In animals with striatonigral lesions, the number of axospinous and axodendritic synapses of the asymmetric type was decreased ipsilateral to the lesions. Synaptic density and ultrastructural integrity remained unaffected in the striata of animals receiving ricin injections and in the contralateral striata of animals receiving OX7-saporin or volkensin injections. Our results, taken together with previous studies showing marked loss of receptors, uptake sites and mRNA, suggest that while most synapses are present and intact, the efficacy of synaptic transmission may be altered.

The Journal of Comparative Neurology, Jan 16, 1995

The m2 muscarinic acetylcholine receptor gene is expressed at high levels in basal forebrain, but... more The m2 muscarinic acetylcholine receptor gene is expressed at high levels in basal forebrain, but the paucity of information about localization of the encoded receptor protein has limited the understanding of cellular and subcellular mechanisms involved in cholinergic actions in this region. The present study sought to determine the cellular localization of m2 protein, its relationship to cholinergic neurons, and its pre-and postsynaptic distribution in the rat medial septum-diagonal band complex using immunocytochemistry with polyclonal rabbit antibodies and a newly developed rat monoclonal antibody specific to the m2 receptor. Light microscopic colocalization studies demonstrated that m2 was present in a subset of choline acetyltransferase immunoreactive neurons, in choline acetyltransferase-negative neurons, and in more neuropil elements than was choline acetyltransferase. Intraventricular injections of 192 IgG-saporin, an immunotoxin directed to the low-affinity nerve growth factor receptor, resulted in depletion of choline acetyltransferase-immunoreactive neurons in the medial septum-diagonal band complex, whereas m2 immunoreactivity in neurons and in the neuropil was unchanged. By electron microscopy, m2 receptor in medial septum-diagonal band complex was localized to the plasmalemma of a small population of small to medium-sized neurons, and it was also found in dendrites, axons, and axon terminals in the neuropil. Neurons expressing m2 immunoreactivity received synaptic contacts from unlabelled axon terminals. A small distinct subpopulation of large neurons, unlabelled by m2 immunoreactivity, received synaptic contacts from m2-immunoreactive terminals. Thus, m2 receptor is situated to mediate the local effects of acetylcholine on basal forebrain cholinergic and noncholinergic neurons and, also, at both pre-and postsynaptic sites.

Journal of Orthopaedic Research, Jul 1, 1997

Neuropathic arthropathy is a severe chronic degenerative condition associated with decreased or a... more Neuropathic arthropathy is a severe chronic degenerative condition associated with decreased or absent sensory innervation of the involved joint. Existing animal models of neuropathic arthritis are limited by the technical difficulties of obtaining either highly selective or complete joint denervation in a minimally invasive fashion. We undertook experiments to determine the feasibility of using the newly described method of selective neuronal lesioning with injected immunotoxin as a means of creating a more tractable model of neuropathic arthritis. Retrograde tracing with fluorochrome revealed that the knee joint of the female Wistar rat is supplied by 581 +/- 31 (mean +/- SD) joint afferents. Immunohistochemistry confirmed that virtually all sensory neurons in the rat express the cell surface receptor Thy 1. Injection of rat knee joints with an immunotoxin targeted toward Thy 1 resulted in the selective ablation of an average of 88% of the joint afferents identified with fluorochrome that are normally found in the ipsilateral L3 and L4 ganglia.

The Journal of Neuroscience the Official Journal of the Society For Neuroscience, Feb 1, 2008

The role of spinal cord-opioid receptor (MOR)-expressing dorsal horn neurons in nociception and m... more The role of spinal cord-opioid receptor (MOR)-expressing dorsal horn neurons in nociception and morphine analgesia is incompletely understood. Using intrathecal dermorphin-saporin (Derm-sap) to selectively destroy MOR-expressing dorsal horn neurons, we sought to determine the role of these neurons in (1) normal baseline reflex nocifensive responses to noxious thermal stimulation (hotplate, tail flick) and to persistent noxious chemical stimulation (formalin) and (2) the antinociceptive activity of intrathecal and systemic morphine in the same tests. Lumbar intrathecal Derm-sap (500 ng) produced (1) partial loss of lamina II MOR-expressing dorsal horn neurons, (2) no effect on MOR-expressing dorsal root ganglion neurons, and (3) no change in baseline tail-flick and hotplate reflex nocifensive responses. Derm-sap treatment attenuated the antinociceptive action of both intrathecal and systemic morphine on hotplate responses. Derm-sap treatment had two effects in the formalin test: (1) increased baseline nocifensive responding and (2) reduced antinociceptive action of systemic morphine. We conclude that MOR-expressing dorsal horn neurons (1) are not essential for determining nocifensive reflex responsiveness to noxious thermal stimuli, (2) are necessary for full antinociceptive action of morphine (intrathecal or systemic) in these tests, and (3) play a significant role in the endogenous modulation of reflex nocifensive responses to persistent pain in the formalin test. Thus, one would predict that altering the activity of MOR-expressing dorsal horn neurons would be antinociceptive and of interest in the search for new approaches to management of chronic pain.

J Neuropathol Exp Neurol, 1993

Journal of Pharmacology and Experimental Therapeutics, Dec 1, 2001

ABSTRACT

Brain Research, Dec 18, 1989

Thy-1 is an abundant surface glycoprotein of rat neurons. OX7 is a monoclonal antibody with high ... more Thy-1 is an abundant surface glycoprotein of rat neurons. OX7 is a monoclonal antibody with high affinity for Thy-1. This study sought to determine if intraventricularly administered OX7 could serve as a carrier to deliver cytotoxin to neurons, thus destroying those neurons. Saporin (Sap), a ribosome-inactivating protein was disulfide-coupled to OX7 (OX7-Sap). OX7-Sap, OX7, saporin alone, pooled non-immune mouse IgG, and an irrelevant immunotoxin, RFT-1-Sap, were injected into the lateral ventricles of anesthetized adult rats. Animals were observed for 1-8 days. OX7-Sap-injected animals developed coarse head tremor and gait/truncal ataxia in a dose-dependent manner beginning 24 h or more after injection. All control animals remained healthy. After OX7 or OX7-Sap injection, immunoperoxidase staining for mouse IgG was most intense and specific in the molecular and Purkinje cell layers of the cerebellar cortex. Cresyl violet staining demonstrated destruction of the Purkinje cell layer in the OX7-Sap-treated animals but not in controls. These results indicate that intraventricular injections of OX7 can be used to deliver biologically active moieties to the Purkinje cells. This approach may prove useful in analysis of Purkinje cell function and as a model of cerebellar degeneration.

Methods in molecular biology (Clifton, N.J.), 2001

ABSTRACT Selective destruction of neurons based on the use of targeted toxins has proven successf... more ABSTRACT Selective destruction of neurons based on the use of targeted toxins has proven successful for several types of neurons (1). This chapter will describe the use of an immunotoxin to selectively destroy rat neurons that express the low-affinity neurotrophin receptor (p75NTR) (2). This immunotoxin consists of a monoclonal antibody disulfide coupled to a ribosome inactivating protein. The most extensively used and studied version uses the antibody 192 IgG originally developed as an antibody to a rat NGF-binding protein (3). 192 IgG has been extensively used to study rat p75NTR. Results of these studies have demonstrated p75NTR expression on a variety of neurotrophin-responsive cells, including sympathetic ganglion neurons, some primary sensory neurons, and cholinergic neurons of the basal forebrain. p75NTR also is expressed on cells not known to be responsive to neurotrophins, such as cerebellar Purkinje neurons and numerous other tissues during development (4). Thus, producing selective lesions using 192 IgG also requires restricting application of the immunotoxin to the region of the target cells.

Neuroscience, 1997

Intracerebroventricular injection of 192 IgG antibody against the p75LNGFR rat low affinity nerve... more Intracerebroventricular injection of 192 IgG antibody against the p75LNGFR rat low affinity nerve growth factor receptor conjugated with saporin, a ribosome inactivating protein, has been shown to destroy the p75LNGFR-expressing cholinergic neurons of the basal forebrain. We injected this immunotoxin into the hippocampus and studied its retrograde effect upon the cholinergic neurons of the medial septum and the vertical limb of the diagonal band of Broca. Seven days after injection, there was a nearly total depletion of cholinergic axons within the hippocampus. This depletion was associated with a marked and significant decrease in the number of cholinergic neurons of the ipsilateral medial septum and the vertical limb of the diagonal band of Broca. At longer survival times, these changes were more pronounced. Parvalbumin-positive, GABAergic neurons within the same areas of the basal forebrain were not affected by immunotoxin injections. Injections of saporin alone had no effect upo...

Cellular and molecular neurobiology, 2003

1. The ability to target specific neurons can be used to produce selective neural lesions and pot... more 1. The ability to target specific neurons can be used to produce selective neural lesions and potentially to deliver therapeutically useful moieties for treatment of disease. In the present study, we sought to determine if a monoclonal antibody to the dopamine transporter (anti-DAT) could be used to target midbrain dopaminergic neurons. 2. The monoclonal antibody recognizes the second, large extracellular loop of DAT. The antibody was conjugated to the "ribosome-inactivating protein"; saporin, and stereotactically pressure microinjected into either the center of the striatum or the left lateral ventricle of adult, male Sprague-Dawley rats. 3. Local intrastriatal injections produced destruction of dopaminergic neurons in the ipsilateral substantia nigra consistent with suicide transport of the immunotoxin. Intraventricular injections (i.c.v.) produced significant loss of dopaminergic neurons in the substantia nigra and ventral tegmental area bilaterally without evident dama...

Neuroscience, 2003

Destruction of neurons in the superficial dorsal horn that express substance P receptor (NK-1R) h... more Destruction of neurons in the superficial dorsal horn that express substance P receptor (NK-1R) has been reported to block development of behavioral hypersensitivity following peripheral sensitization of nociceptors. Baseline sensitivity was not altered in these rat models that assessed innate reflex responses (i.e. hind-paw withdrawal to thermal or mechanical stimulation). In the present study, we evaluated effects of intrathecal substance P-saporin (SP-sap), a toxin selective for cells expressing NK-1R, on operant escape responses of rats to thermal stimulation. For comparison, lick/guard reflex testing was performed. Injection of a modest dose (175 ng) of SP-sap into the lumbar subarachnoid space produced a partial loss of lamina I/II NK-1R-expressing dorsal horn neurons but did not affect NK-1R-expressing neurons in deeper laminae. Lick/guard responses to 0.3, 44 or 47 degrees C were not affected after SP-sap treatment, but escape responses to these temperatures were significant...