The microglial-motoneuron dialogue in ALS.

Neuroinflammation is a pathological hallmark of neurodegenerative diseases including amyotrophic lateral sclerosis (ALS), and is characterized by activated microglia at sites of neuronal injury. In ALS, neurons do not die alone; neuronal injury is noncell-autonomous and depends upon a well-orchestrated dialogue between motor neurons and microglia. Evidence from transgenic models expressing mutant superoxide dismutase 1 (SOD) suggests that the dialogue between motor neurons and microglia initially protects motor neurons. However, with increasing stress and injury within motor neurons, induced by the presence of misfolded proteins such as mSOD1, mitochondrial function and axoplasmic flow are impaired and endoplasmic reticulum stress is induced; misfolded proteins themselves or alternate signals are released from motor neurons and activate microglia. Activated microglia, in turn, switch from anti-inflammatory and neuroprotective to proinflammatory and neurotoxic. Neurotoxic signaling from motor neurons promotes microglial release of reactive oxygen species and pro-inflammatory cytokines further enhancing motor neuron stress and cell injury and initiating a self-propagating cycle of motor neuron injury and cell death. A greater understanding of how to restore the imbalance between neuroprotection and cytotoxicity will depend upon a greater understanding of the motor neuron-microglial dialogue.

Neuropathology of herpes simplex virus encephalitis in a rat seizure model.

Herpes simplex virus type 1 (HSV-1) is the cause of a serious and often fatal encephalitis. Patients who survive herpes simplex encephalitis (HSE) experience behavioral abnormalities including profound cognitive dysfunctions. We have developed a rat model of acute HSE to investigate the cognitive impairments caused by HSV-1 central nervous system (CNS) infection. Following intranasal inoculation of Lewis rats with a neurovirulent strain of HSV-1, animals shed virus in both ocular and nasal secretions and developed clinical signs of infection, including partial complex motor seizures that eventually generalized. Homogenization assays demonstrated infectious virus in the trigeminal ganglia, olfactory bulbs, and the piriform and entorhinal cortices. Histopathological assessment revealed inflammatory and hemorrhagic lesions in the trigeminal ganglia, olfactory bulbs, amygdala, hippocampus, the piriform and entorhinal cortices, and the spinal trigeminal nuclei. Viral antigens and nucleic acids were also detected within these structures by immunofluorescence microscopy and in situ hybridization, respectively. Viral-induced astrocytic hypertrophy in the CNS was demonstrated by glial fibrillary acidic protein immunoreactivity. Together, these results indicate that HSV-1 has the ability to invade, replicate, and induce site-specific CNS damage in the Lewis rat.

Serial position curves for item (spatial location) information: role of the dorsal hippocampal formation and medial septum.

Animals were trained on an item recognition memory task for a list of 5 spatial locations. After reaching criterion performance the animals sustained small or medium-size dorsal hippocampal formation lesions, small or large medial septal lesions, or served as sham-operated or cortical controls. Following recovery from surgery, animals were retested for item recognition memory. Sham-operated and cortical control animals showed no deficits in performance. In contrast, animals with small dorsal hippocampal formation or medial septal lesions displayed a deficit for the early items, but had excellent memory for the last item of the list. Animals with medium-size dorsal hippocampal formation or large medial septal lesions displayed a deficit for both early and late items within the list. Because residual short-term memory capacity can be seen only with small hippocampal formation or medial septal lesions, it is suggested that the hippocampal formation and cholinergic input into the hippocampal formation via the medial septum code spatial information within a continuous extended time frame.

Spatial recognition memory deficits without notable CNS pathology in rats following herpes simplex encephalitis.

Survivors of herpes simplex encephalitis (HSE) experience intellectual impairment and an inability to store and recall information. Because the temporal lobes and associated limbic structures are central to storage and retrieval of memories, and are predominantly affected in adult HSE, injury to these areas is postulated to cause behavioral and learning disabilities. A previous study (Beers et al., 1993) demonstrated that intranasal inoculation of Lewis rats with herpes simplex virus type-1 (HSV-1) induced acute partial complex seizures, and hemorrhagic and inflammatory lesions of the hippocampus and entorhinal cortex. Consequently, it was of interest to determine whether rats that had recovered from HSE had limbic system-associated memory impairments. Therefore, rats were evaluated when signs and symptoms of encephalitis were no longer apparent using an eight arm radial maze to assess the acquisition and retention of learned information. An allocentric-spatial location paradigm revealed HSV-1 infected rats performed at chance levels on both acquisition and retention which were statistically different from sham-inoculated controls. However, using an egocentric-spatial left/right discrimination task, infected rats performed statistically similar to sham-inoculated controls. Furthermore, HSV-1 nucleic acids were detected in the nuclei of neurons within the hippocampus and entorhinal cortex using in situ hybridization techniques. Of interest was the observation that rats with learning and memory deficits had no apparent histopathological or immunocytochemical evidence of antecedent CNS infection. This is the first experimental demonstration that HSV-1 can cause behavioral impairments in the absence of obvious inflammatory injury to the temporal lobe memory system.

Hematopoietic stem cell transplantation in patients with sporadic amyotrophic lateral sclerosis.

BACKGROUND: Amyotrophic lateral sclerosis (ALS), an inexorably progressive motoneuron disease, is accompanied by significantly increased markers of inflammation. These inflammatory constituents could protect, harm, do neither, or do both. OBJECTIVE: Allogeneic hematopoietic stem cell transplantation (HSCT) was performed in patients with sporadic ALS to suppress neuroinflammation and improve clinical outcomes after CNS engraftment. METHODS: Six patients with definite ALS received total body irradiation followed by peripheral blood HSCT infusion from human leukocyte antigen identically matched sibling donors. Disease progression and survival were assessed monthly and compared with matched historic database patients. Autopsy samples from brain and spinal cord were examined immunohistochemically and by quantitative reverse-transcriptase polymerase chain reaction. Donor-derived DNA in brain and spinal cord tissue was evaluated for the extent of chimerism. RESULTS: No clinical benefits were evident. Four patients were 100% engrafted; postmortem tissue examination in two of the 100% engrafted patients demonstrated 16% to 38% donor-derived DNA at sites with motoneuron pathology, which may correspond to the observed increased CD68 or CD1a-positive cells. Neither donor DNA nor increased cell numbers were found in several unaffected brain regions. A third minimally engrafted patient had neither donor DNA nor increased infiltrating cells in the CNS. CONCLUSIONS: This study demonstrates that peripheral cells derived from donor hematopoietic stem cells can enter the human CNS primarily at sites of motoneuron pathology and engraft as immunomodulatory cells. Although unmodified hematopoietic stem cells did not benefit these sporadic amyotrophic lateral sclerosis patients, such cells may provide a cellular vehicle for future CNS gene therapy.

Dissociation of data-based and expectancy-based memory following hippocampal lesions in rats.

Sham-operated and nonoperated animals or animals with hippocampal lesions were presented with sets of trials to test both expectancy-based and data-based memory within the same task. During the study phase of each trial the animals were presented with a constant sequence of five arms on an eight-arm radial maze followed by a test phase in which a recognition test requiring a win-stay rule was used. Expectancy-based memory was measured during the study phase of the trials as a pattern of correct or incorrect orienting responses in anticipation of the ensuing doors in the constant sequence. Both groups of animals acquired correct orienting responses at the same rate, emitted the same pattern of correct orienting responses, and made the same number and pattern of intralist and extralist intrusion errors. Data-based memory was measured during the test phase of the trial as correct recognition test performance. During the test phase the animals with hippocampal lesions were impaired relative to controls on both immediate and 24-h recognition tests. These results suggest that the hippocampus might mediate only data-based, but not expectancy-based, memory and imply a possible dissociation between expectancy-based and data-based memory systems.

HFE mutations are not strongly associated with sporadic ALS.

The presence of oxidative damage and increased iron deposition in CNS tissues of ALS patients prompted the authors to examine the prevalence of two common HFE gene mutations linked to iron accumulation and consequent oxidative stress. The prevalence of the C282Y and H63D mutations was nearly identical in 51 ALS patients and 47 normal control subjects. The presence of either mutation did not significantly affect the age at onset or rate of progression in ALS.

Role of potassium channels in amyloid-induced cell death.

Basal forebrain cholinergic neurons are severely depleted early in Alzheimer's disease and appear particularly susceptible to amyloid beta-peptide (A beta) toxicity in vivo. To model this effect in vitro, a cholinergic septal cell line (SN56) was exposed to A beta. SN56 cells exhibited a tetraethylammonium (TEA)-sensitive outward K+ current with delayed rectifier characteristics. Increases of 64% (+/-19; p < 0.02) and 44% (+/-12; p < 0.02) in K+ current density were noted 6-12 and 12-18 h following the addition of A beta to SN56 cell cultures, respectively. Morphological observation and staining for cell viability showed that 25 +/- 4 and 39 +/- 4% of SN56 cells were dead after 48- and 96-h exposures to A beta, respectively. Perfusion of SN56 cells with 10-20 mM TEA blocked 71 +/- 6 to 92 +/- 2% of the outward currents, widened action potentials, elevated [Ca2+]i, and inhibited 89 +/- 14 and 68 +/- 14% of the A beta toxicity. High [K+]o, which depolarizes cell membranes and increases [Ca2+]i, also protected SN56 cells from A beta toxicity. This effect appeared specific since glucose deprivation of SN56 cells did not alter K+ current density and TEA did not protect these cells from hypoglycemic cell death. Furthermore, A beta was toxic to a dopaminergic cell line (MES23.5) that expressed a K+ current with delayed rectifier characteristics; K+ current density was not altered by A beta and MES23.5 cells were not protected by TEA from A beta toxicity. In contrast, a noncholinergic septal cell line (SN48) that shows minimal outward K+ currents was resistant to the toxicity of A beta. These data suggest that a K+ channel with delayed rectifier characteristics may play an important role in A beta-mediated toxicity for septal cholinergic cells.

Cloning and restriction endonuclease mapping of herpes simplex virus type-1 strains H129 and +GC.

EcoRI fragments of herpes simplex virus I (HSV-1) strains H129 and +GC were cloned and the EcoRI and BglII restriction enzyme sites were mapped. Comparison of these enzyme sites with the sequence of HSV-1 strain 17syn+ demonstrated that all EcoRI sites were identical. For H129, the BglII sites were also found to match strain 17syn+ BglII sites. With one exception, the BglII sites in strain +GC also aligned with the strain 17syn+ sequence. The one exception was a missing BglII site from strain +GC located between bases 25,149 and 25,154 in the EcoRI D fragment within the viral deoxyribonuclease gene (UL12). The BglII site represents the first difference to be mapped within HSV-1 strains H129 and +GC which have unique pathobiological properties in animal models of acute and reactivated infections.

Parvalbumin overexpression alters immune-mediated increases in intracellular calcium, and delays disease onset in a transgenic model of familial amyotrophic lateral sclerosis.

Intracellular calcium is increased in vulnerable spinal motoneurons in immune-mediated as well as transgenic models of amyotrophic lateral sclerosis (ALS). To determine whether intracellular calcium levels are influenced by the calcium-binding protein parvalbumin, we developed transgenic mice overexpressing parvalbumin in spinal motoneurons. ALS immunoglobulins increased intracellular calcium and spontaneous transmitter release at motoneuron terminals in control animals, but not in parvalbumin overexpressing transgenic mice. Parvalbumin transgenic mice interbred with mutant SOD1 (mSOD1) transgenic mice, an animal model of familial ALS, had significantly reduced motoneuron loss, and had delayed disease onset (17%) and prolonged survival (11%) when compared with mice with only the mSOD1 transgene. These results affirm the importance of the calcium binding protein parvalbumin in altering calcium homeostasis in motoneurons. The increased motoneuron parvalbumin can significantly attenuate the immune-mediated increases in calcium and to a lesser extent compensate for the mSOD1-mediated 'toxic-gain-of-function' in transgenic mice.