Timothy E. O'Brien

Functional recovery and neuroanatomical plasticity following middle cerebral artery occlusion and IN-1 antibody treatment in the adult rat

Stroke is a prevalent and devastating disorder, and no treatment is currently available to restore lost neuronal function after stroke occurs. One unique therapy that may improve functional recovery after stroke is blockade of the neurite inhibitory protein Nogo‐A with the monoclonal antibody IN‐1, through enhancement of neuroanatomical plasticity from uninjured areas of the central nervous system. In the present study, we combined IN‐1 treatment with an ischemic lesion (permanent middle cerebral artery occlusion) to determine the effect of Nogo‐A neutralization on cortical plasticity and functional recovery. We report here that, following ischemic stroke and treatment with IN‐1, adult rats demonstrated functional recovery on a forelimb‐reaching task and new cortico‐efferent projections from the opposite, unlesioned hemisphere. These results support the efficacy of Nogo‐A blockade as a treatment for ischemic stroke and implicate plasticity from the unlesioned hemisphere as a mechanism for recovery.

Delayed Treatment with Monoclonal Antibody IN-1 1 Week after Stroke Results in Recovery of Function and Corticorubral Plasticity in Adult Rats

Neuronal death due to ischemic stroke results in permanent deficits in sensory, language, and motor functions. The growth-restrictive environment of the adult central nervous system (CNS) is an obstacle to functional recovery after stroke and other CNS injuries. In this regard, Nogo-A is a potent neurite growth-inhibitory protein known to restrict neuronal plasticity in adults. Previously, we have found that treatment with monoclonal antibody (mAb) IN-1 to neutralize Nogo-A immediately after stroke enhanced motor cortico-efferent plasticity and recovery of skilled forelimb function in rats. However, immediate treatment for stroke is often not clinically feasible. Thus, the present study was undertaken to determine whether cortico-efferent plasticity and functional recovery would occur if treatment with mAb IN-1 was delayed 1 week after stroke. Adult rats were trained on a forelimb-reaching task, and the middle cerebral artery was occluded to induce focal cerebral ischemia to the forelimb sensorimotor cortex. After 1 week, animals received mAb IN-1 treatment, control antibody, or no treatment, and were tested for 9 more weeks. To assess cortico-efferent plasticity, the sensorimotor cortex opposite the stroke lesion was injected with an anterograde neuroanatomical tracer. Behavioral analysis demonstrated a recovery of skilled forelimb function, and anatomical studies revealed neuroplasticity at the level of the red nucleus in animals treated with mAb IN-1, thus demonstrating the efficacy of this treatment even if administered 1 week after stroke.

Recovery and brain reorganization after stroke in adult and aged rats

Stroke is a prevalent and devastating disorder, and no treatment is currently available to restore lost neuronal function after stroke. One unique therapy that improves recovery after stroke is neutralization of the neurite inhibitory protein Nogo‐A. Here, we show, in a clinically relevant model, improved functional recovery and brain reorganization in the aged and adult rat when delayed anti–Nogo‐A therapy is given after ischemic injury. These results support the efficacy of Nogo‐A neutralization as treatment for ischemic stroke, even in the aged animal and after a 1‐week delay, and implicate neuronal plasticity from unlesioned areas of the central nervous system as a mechanism for recovery. Ann Neurol 2005;58:950–953

Axonal plasticity is associated with motor recovery following amphetamine treatment combined with rehabilitation after brain injury in the adult rat

Clinical and laboratory studies have suggested that amphetamine treatment when paired with rehabilitation results in improved recovery of function after stroke or traumatic brain injury. In the present study, we investigated whether new anatomical pathways developed in association with improved motor function after brain damage and amphetamine treatment linked with rehabilitation. Following a unilateral sensorimotor cortex lesion in the adult rat, amphetamine (2 mg/kg) was administered in conjunction with physiotherapy sessions on postoperative days two and five. Physiotherapy was continued twice daily for the first 3 weeks after injury, and then once daily until week six. Performance on skilled forelimb reaching and ladder rung walking was used to assess motor improvement. Our results show that animals with sensorimotor cortical lesions receiving amphetamine treatment linked with rehabilitation had significant improvement in both tasks. Neuroanatomical tracing of efferent pathways from the opposite, non-damaged cortex resulted in the novel finding that amphetamine treatment linked with rehabilitation, significantly increased axonal growth in the deafferented basilar pontine nuclei. These results support the notion that pharmacological interventions paired with rehabilitation can enhance neuronal plasticity and thereby improve functional recovery after CNS injury.

Mutation in murine coronavirus replication protein nsp4 alters assembly of double membrane vesicles

Abstract Coronaviruses are positive-strand RNA viruses that replicate in the cytoplasm of infected cells by generating a membrane-associated replicase complex. The replicase complex assembles on double membrane vesicles (DMVs). Here, we studied the role of a putative replicase anchor, nonstructural protein 4 (nsp4), in the assembly of murine coronavirus DMVs. We used reverse genetics to generate infectious clone viruses (icv) with an alanine substitution at nsp4 glycosylation site N176 or N237, or an asparagine to threonine substitution (nsp4-N258T), which is proposed to confer a temperature sensitive phenotype. We found that nsp4-N237A is lethal and nsp4-N258T generated a virus (designated Alb ts6 icv) that is temperature sensitive for viral replication. Analysis of Alb ts6 icv-infected cells revealed that there was a dramatic reduction in DMVs and that both nsp4 and nsp3 partially localized to mitochondria when cells were incubated at the non-permissive temperature. These results reveal a critical role of nsp4 in directing coronavirus DMV assembly.

Effects of mixtures of phenolic acids on phosphorus uptake by cucumber seedlings

To determine how individual phenolic acids in a mixture might affect phosphorus (P) uptake, 15-day-old cucumber seedlings grown in solution culture were treated with ferulic, vanillic,p-coumaric, or equimolar mixtures of these phenolic acids. Phenolic acid and P uptake were determined by solution depletion. The joint action of the mixtures of these phenolic acids on P uptake was primarily additive. Thus, as the number of phenolic acids increased in the mixture, the concentrations of the individual phenolic acids in the mixture required to bring about a given response declined. Seedling uptake of individual phenolic acids from solution mixtures of phenolic acids was reduced when compared to the uptake of phenolic acids from single phenolic acid solutions. The magnitude of the reduction varied with phenolic acid and concentration. The dose required for 50% inhibition of P uptake was approximately two to three times higher for vanillic acid (6.73 mM) than for ferulic (2.27 mM) andp-coumaric acids (3.00 mM) when dose was based on the initial treatment concentrations. The dose required for 50% inhibition of P uptake was not significantly different for the three phenolic acids (42 ± 5 μmol/g root fresh weight) when dose was based on phenolic acid uptake. Potential reasons for these differences are discussed.

A Gentle Introduction to Optimal Design for Regression Models

This article demonstrates and underscores the equivalence between a variance-maximization exercise and the methodology involved in obtaining and verifying the optimal design for a key model function. It thus provides an alternate solution to the variance exercise as well as a means to introduce and illustrate the concepts of optimal design theory and practice in a simple and clear manner.

Microglia play a role in mediating the effects of cytokines on the structure and function of the rat pineal gland

Abstract. The role of the pineal gland in regulating immune function has been extensively investigated. However, there is little information about possible feedback mechanisms of immunological factors on pineal gland neuroendocrine functions. Therefore, experiments were designed to test the effects of cytokines (interferon-gamma, IFN-γ; interleukin-1β, IL-1β; tumor necrosis factor-alpha, TNF-α; transforming growth factor-β1, TGF-β1) on pinealocytes and the role of pineal microglia in mediating these cytokine effects in the pineal gland of the rat. Our studies showed that IFN-γ enhanced 5-hydroxytryptamine (5-HT) content (measured by high-performance liquid chromatography, HPLC) and increased pinealocyte process length in pineal cultures. IL-1β treatment decreased 5-HT content in both cell and organ culture, but exhibited no effect on pinealocyte process length. 5-HT content and process length were decreased by TNF-α treatment. IFN-γ and IL-1β exhibited no significant effect in the absence of microglia in cell cultures. In contrast, TNF-α caused a further decline in 5-HT content even in the absence of microglia in the cultures. The effects of TNF-α were probably due to toxic effects, since an increased number of pyknotic nuclei were observed in treated cultured explants. TGF-β1 treatment caused aggregation of pinealocytes in cultures and suppressed process length and 5-HT content. In conclusion, cytokine effects on pinealocytes may be mediated by microglia (IFN-γ and IL-1β) or act directly on pinealocytes (TNF-α). The presence of IL-1β and TGF-β1 protein in the pineal gland and the suppressive effect of TGF-β1 on pinealocytes in cultures further suggest that endogenous cytokines play regulatory roles in response to peripheral homeostatic changes.

Model-robust and model-sensitive designs

The main drawback of the optimal design approach is that it assumes the statistical model is known. To overcome this problem, a new approach to reduce the dependency on the assumed model is proposed. The approach takes into account the model uncertainty by incorporating the bias in the design criterion and the ability to test for lack-of-fit. Several new designs are derived and compared to the alternatives available from the literature.

Anti-Nogo-A Immunotherapy Does Not Alter Hippocampal Neurogenesis after Stroke in Adult Rats

Ischemic stroke is a leading cause of adult disability, including cognitive impairment. Our laboratory has previously shown that treatment with function-blocking antibodies against the neurite growth inhibitory protein Nogo-A promotes functional recovery after stroke in adult and aged rats, including enhancing spatial memory performance, for which the hippocampus is critically important. Since spatial memory has been linked to hippocampal neurogenesis, we investigated whether anti-Nogo-A treatment increases hippocampal neurogenesis after stroke. Adult rats were subject to permanent middle cerebral artery occlusion followed 1 week later by 2 weeks of antibody treatment. Cellular proliferation in the dentate gyrus was quantified at the end of treatment, and the number of newborn neurons was determined at 8 weeks post-stroke. Treatment with both anti-Nogo-A and control antibodies stimulated the accumulation of new microglia/macrophages in the dentate granule cell layer, but neither treatment increased cellular proliferation or the number of newborn neurons above stroke-only levels. These results suggest that anti-Nogo-A immunotherapy does not increase post-stroke hippocampal neurogenesis.