Transplanted Human Neural Progenitor Cells Attenuate Motor Dysfunction and Lengthen Longevity in a Rat Model of Ataxia.

The spastic Han Wistar (sHW) rat serves as a model for human ataxia presenting symptoms of motor deterioration, weight loss, shortened lifespan, and Purkinje neuron loss. Past studies revealed that human neural progenitor cells (NPCs) improved ataxic symptoms at 20 d posttransplantation in sHW rats. In this study, we investigated the fate and longer-term effectiveness of these transplanted NPCs. Rats were placed into four treatment groups: an untreated normal control group (n = 10), an untreated mutant rat control (n = 10), a mutant group that received an injection of dead NPCs (n = 9), and a mutant group that received live NPCs (n = 10). Bilateral cerebellar injections containing 500,000 of either live or dead NPCs were performed on mutant sHW rats at 40 d of age. Motor activity for all mutant rats started to decline in open field testing around day 35. However, at day 45, the live NPC-treated mutants exhibited significant improvements in open field activity. Similar improvements were observed during rotarod testing and weight gain through the completion of the experiments (100 d). Immunohistochemistry revealed few surviving human NPCs in the cerebella of 80- and 100-d-old NPC-treated mutants; while cresyl violet staining revealed that live NPC-treated mutants had significantly more surviving Purkinje neurons compared to mutants that were untreated or received dead NPCs. Direct stereotactic implantation of NPCs alleviated the symptoms of ataxia, acting as a neuroprotectant, supporting future clinical applications of these NPCs in the areas of ataxia as well as other neurodegenerative diseases.

Induced expression of a vestigial sexual signal.

Vestigial morphological traits are common and well known in a variety of taxa. Identification of vestigial genes has illustrated the potential for evolutionary reversals and the re-expression of atavistic traits. Here we induce expression of a behavioural sexual signal, male calling song, in a cricket species, Gryllus ovisopis, which lacks a functional calling song. We successfully used acetylcholine injections in the frontal space of the head of male crickets to activate cerebral command neurons for cricket calling, and we recorded calling songs with a temporal chirp pattern similar to that of G. ovisopis' close evolutionary relatives, G. firmus and G. pennsylvanicus, implying that the neural pattern generators that underlie cricket calling behaviour persist in a vestigial state in G. ovisopis To our knowledge, this is the first demonstration of the induced expression of a vestigial behaviour in any organism. The retention of latent neural capacity to express sexual behaviours could have important implications for rapid evolution, trait re-emergence and reproductive isolation.

Validation of Acoustic Wave Induced Traumatic Brain Injury in Rats.

BACKGROUND: This study looked to validate the acoustic wave technology of the Storz-D-Actor that inflicted a consistent closed-head, traumatic brain injury (TBI) in rats. We studied a range of single pulse pressures administered to the rats and observed the resulting decline in motor skills and memory. Histology was observed to measure and confirm the injury insult. METHODS: Four different acoustic wave pressures were studied using a single pulse: 0, 3.4, 4.2 and 5.0 bar (n = 10 rats per treatment group). The pulse was administered to the left frontal cortex. Rotarod tests were used to monitor the rats' motor skills while the water maze test was used to monitor memory deficits. The rats were then sacrificed ten days post-treatment for histological analysis of TBI infarct size. RESULTS: The behavioral tests showed that acoustic wave technology administered an effective insult causing significant decreases in motor abilities and memory. Histology showed dose-dependent damage to the cortex infarct areas only. CONCLUSIONS: This study illustrates that the Storz D-Actor effectively induces a repeatable TBI infarct, avoiding the invasive procedure of a craniotomy often used in TBI research.

Efficacy of Two Delivery Routes for Transplanting Human Neural Progenitor Cells (NPCs) Into the Spastic Han-Wistar Rat, a Model of Ataxia.

An emerging avenue for recalcitrant neurodegenerative disease treatment is neural progenitor cell (NPC) transplantation. In this study, we investigated the effectiveness of two different delivery routes of human-derived NPC inoculation: injection into the common carotid artery or unilateral stereotactic implantation into the degenerating cerebellum and hippocampus of spastic Han-Wistar (sHW) rats, a model of ataxia. At 30 days of age, sHW mutants were implanted with osmotic pumps preloaded with cyclosporine. Ten days after pump implantation, the animals were given either 3,000,000 live human-derived NPCs (hNPCs; n = 12) or 3,000,000 dead NPCs (dNPCs; n = 12) injected into the common carotid artery, or were given two unilateral implantations of 500,000 hNPCs into the cerebellum and 500,000 hNPCs into the hippocampus of each sHW rat (n = 12) or 500,000 dNPCs by unilateral implantation into the cerebellum and hippocampus (n = 12). We also compared treated sHW rats to untreated sHW rats: normal rats (n = 12) and sibling sHW rats (n = 12). Motor activity and animal weights were monitored every 5 days to ascertain effectiveness of the two types of delivery methods compared to the untreated mutant and normal animals. Mutant rats with hNPC implantations, but not dNPC or carotid artery injections, showed significant deceleration of motor deterioration (p < 0.05). These mutants with hNPC implantations also retained weight longer than dNPC mutants did (p < 0.05). At the end of the experiment, animals were sacrificed for histological evaluation. Using fluorescent markers (Qtracker) incorporated into the hNPC prior to implantation and human nuclear immunostaining, we observed few hNPCs in the brains of carotid artery-injected mutants. However, significant numbers of surviving hNPCs were seen using these techniques in mutant cerebellums and hippocampi implanted with hNPC. Our results show that direct implantation of hNPCs reduced ataxic symptoms in the sHW rat, demonstrating that stereotactic route of stem cell delivery correlates to improved clinical outcomes.

Transplantation of Human Neural Progenitor Cells Reveals Structural and Functional Improvements in the Spastic Han-Wistar Rat Model of Ataxia.

The use of regenerative medicine to treat nervous system disorders like ataxia has been proposed to either replace or support degenerating neurons. In this study, we assessed the ability of human neural progenitor cells (hNPCs) to repair and restore the function of dying neurons within the spastic Han-Wistar rat (sHW), a model of ataxia. The sHW rat suffers from neurodegeneration of specific neurons, including cerebellar Purkinje cells and hippocampal CA3 pyramidal cells leading to the observed symptoms of forelimb tremor, hind-leg rigidity, gait abnormality, motor incoordination, and a shortened life span. To alleviate the symptoms of neurodegeneration and to replace or augment dying neurons, neuronal human progenitor cells were implanted into the sHW rats. At 30 d of age, male sHW mutant rats underwent subcutaneous implantation of an Alzet osmotic pump that infused cyclosporine (15 mg/kg/d) used to suppress the rat's immune system. At 40 d, sHW rats received bilateral injections (500,000 cells in 5 µL media) of live hNPCs, dead hNPCs, live human embryonic kidney cells, or growth media either into the cerebellar cortex or into the hippocampus. To monitor results, motor activity scores (open-field testing) and weights of the animals were recorded weekly. The sHW rats that received hNPC transplantation into the cerebellum, at 60 d of age, displayed significantly higher motor activity scores and sustained greater weights and longevities than control-treated sHW rats or any hippocampal treatment group. In addition, cerebellar histology revealed that the transplanted hNPCs displayed signs of migration and signs of neuronal development in the degenerated Purkinje cell layer. This study revealed that implanted human progenitor cells reduced the ataxic symptoms in the sHW rat, identifying a future clinical use of these progenitor cells against ataxia and associated neurodegenerative diseases.

Exercise-induced neuroprotection in the spastic Han Wistar rat: the possible role of brain-derived neurotrophic factor.

Moderate aerobic exercise has been shown to enhance motor skills and protect the nervous system from neurodegenerative diseases, like ataxia. Our lab uses the spastic Han Wistar rat as a model of ataxia. Mutant rats develop forelimb tremor and hind limb rigidity and have a decreased lifespan. Our lab has shown that exercise reduced Purkinje cell degeneration and delayed motor dysfunction, significantly increasing lifespan. Our study investigated how moderate exercise may mediate neuroprotection by analyzing brain-derived neurotrophic factor (BDNF) and its receptor TrkB. To link BDNF to exercise-induced neuroprotection, mutant and normal rats were infused with the TrkB antagonist K252a or vehicle into the third ventricle. During infusion, rats were subjected to moderate exercise regimens on a treadmill. Exercised mutants receiving K252a exhibited a 21.4% loss in Purkinje cells compared to their controls. Cerebellar TrkB expression was evaluated using non-drug-treated mutants subjected to various treadmill running regimens. Running animals expressed three times more TrkB than sedentary animals. BDNF was quantified via Sandwich ELISA, and cerebellar expression was found to be 26.6% greater in mutant rats on 7-day treadmill exercise regimen compared to 30 days of treadmill exercise. These results suggest that BDNF is involved in mediating exercise-induced neuroprotection.

Muscle-specific changes in length-force characteristics of the calf muscles in the spastic Han-Wistar rat.

The purpose of the present study was to investigate muscle mechanical properties and mechanical interaction between muscles in the lower hindlimb of the spastic mutant rat. Length-force characteristics of gastrocnemius (GA), soleus (SO), and plantaris (PL) were assessed in anesthetized spastic and normally developed Han-Wistar rats. In addition, the extent of epimuscular myofascial force transmission between synergistic GA, SO, and PL, as well as between the calf muscles and antagonistic tibialis anterior (TA), was investigated. Active length-force curves of spastic GA and PL were narrower with a reduced maximal active force. In contrast, active length-force characteristics of spastic SO were similar to those of controls. In reference position (90° ankle and knee angle), higher resistance to ankle dorsiflexion and increased passive stiffness was found for the spastic calf muscle group. At optimum length, passive stiffness and passive force of spastic GA were decreased, whereas those of spastic SO were increased. No mechanical interaction between the calf muscles and TA was found. As GA was lengthened, force from SO and PL declined despite a constant muscle-tendon unit length of SO and PL. However, the extent of this interaction was not different in spastic rats. In conclusion, the effects of spasticity on length-force characteristics were muscle specific. The changes observed for GA and PL muscles are consistent with the changes in limb mechanics reported for human patients. Our results indicate that altered mechanics in spastic rats cannot be attributed to differences in mechanical interaction, but originate from individual muscular structures.

Dopamine as an anorectic neuromodulator in the cockroach Rhyparobia maderae.

Insects, including cockroaches, self-select a balanced diet when faced with different nutrient choices. For self-selection to be carried out effectively, insects possess neuroregulatory systems to control their food intake. In the present study, we examined the role of the neurotransmitter dopamine (DA) in the feeding regulation of the Madeira cockroach (Rhyparobia maderae). When R. maderae nymphs were injected with 20 µl of 100 mmol l(-1) DA, they showed an 83.3% reduction in sucrose intake and a 78.9% reduction in total intake compared with saline-injected controls. The DA agonist, 2-amino-6,7-dihydroxy-1,2,3,4-tetrahydronaphthalene (6,7-ADTN) (100 mmol l(-1) in 1 µl), caused a significant reduction in sucrose feeding, reducing feeding by 47.3% compared with saline-injected controls. Protein feeding was also significantly reduced by 6,7-ADTN to 62%. Rhyparobia maderae nymphs injected with the DA antagonist chlorpromazine (100 mmol l(-1) in 1 µl) did not differ significantly from control nymphs in their feeding behavior. Interestingly, R. maderae nymphs injected with 2 µl or 5 µl chlorpromazine (100 mmol l(-1)) showed significantly increased mortality rates of 47.5% or 66.7%, respectively. The DA antagonist, spiperone (100 mmol l(-1) in 1 µl), caused a significant feeding response, showing an increase in feeding in both sucrose (310.6%) and total intake (236.3%). Casein feeding in R. maderae nymphs was also elevated (70.8%) but this was not statistically significant. The experiments with DA, the DA agonist 6,7-ADTN and the DA antagonist spiperone strongly suggest that the neurotransmitter DA is involved in regulating feeding in the cockroach R. maderae.

Neuroprotective effects of moderate aerobic exercise on the spastic Han-Wistar rat, a model of ataxia.

Research has shown that physical exercise may reduce degeneration in certain brain regions experiencing ataxia. Our laboratory utilized mutant spastic Han-Wistar rats (sHW) that display developmental abnormalities, including spastic paresis, fore limb tremors, hind limb rigidity, and a reduced life span (60-65 days of age). Concomitant neurodegeneration has been observed in the cerebellum (Purkinje cells). The purpose of this study was to investigate if moderate, aerobic exercise could reduce Purkinje cell neurodegeneration and improve the motor ability and survival of the mutant sHW rat. Mutant male littermates at the ages of 20 (n=11 pairs) and 30 (n=13 pairs) days old were divided into running groups and non-running groups. Mutant rats were run on a motorized treadmill at the rate of 15 m/min with a 10% slope. The "running" group ran for 30 min per day, 5 days a week; the "non-runners" remained nearby in the training facility. These conditions were held constant until the mutant runners could no longer run due to disease progression. Moderate exercise increased the lifespan of running mutant rats in both the 20-day start group (14% increase) and 30-day start group (13% increase). The rats exhibited improved motor function as open-field tests showed higher activity scores for runners after 50 days. Histological examination of the cerebellum revealed a 62% increase in Purkinje cell survival of the runners. These results suggest that aerobic exercise ameliorates, at least partially, cerebellar dysfunction in the sHW rat, an excellent model of ataxia.

Phototactic responses to ultraviolet and white light in various species of Collembola, including the eyeless species, Folsomia candida.

Previous observations have indicated homology in the cellular components between Collembolan eyes and the compound eyes of insects. However, behavioral or physiological studies indicating similarities in function are lacking. Collembolan eyes were examined from three species in the Family Isotomidae using scanning electron microscopy. Collembolan eyes are arranged dorsally and laterally on each side of the head in two species, Proisotoma minuta with eight eyes on each side of the head and Folsomia similis with one eye on each side of the head. In both of these species the eyes were located just posterior to the postantennal organ. In Folsomia candida, no external eye structures were detected. These three species were assayed for a series of behavioral preferences using ultraviolet (UV), white light and dark, and temperature conditions. The tests demonstrated that over 76% of all three species, including the eyeless F. Candida, chose white over UV light, over 69% preferred dark over UV, and over 77% favored dark over white light. The results demonstrated that all three species detect both UV and white light and avoid it, preferring cool, dark habitats. From the results of this study, it is hypothesized that F. candida may, in fact, be only "lensless" and may be able to detect light by having internal, non-ocular photoreceptors. Further histological studies are needed to investigate this possibility.

Characterization of chronic nicotine exposure on the survival of the spastic Han-Wistar rat.

Excitotoxicity has been implicated as a mechanism of cell death in many neurodegenerative disorders. Cell culture studies have shown that neuroprotection can be induced by preincubation with the acetylcholine agonist nicotine. We investigated the possible neuroprotective effects of nicotine in the spastic Han-Wistar rat, which suffers from glutamate excitotoxicity affecting two central nervous system regions: The hippocampus and the cerebellum. To investigate nicotine's possible neuroprotection, we treated 25-day-old mutant and normal siblings with 50-75 mg/l nicotine in their drinking solutions. The 75-ml/l dose significantly improved motor activity and increased longevity of the mutants (p<.05). To assess whether nicotine protected individual neurons, we performed hematoxylin and eosin (H&E) staining of brain sections. The histological data indicated that nicotine increased the survival of Purkinje cells in the mutants by as much as 50% but did not prevent cell death. To investigate whether the neuroprotection by nicotine was due to changes in nicotinic receptor expression, we performed immunohistochemical studies by staining for the alpha 3, alpha 4, and alpha 7 receptor subunits in mutant and normal rats. The alpha 4 subunit was upregulated by nicotine treatment in the cerebellum and was noted to have lower levels throughout the hippocampus of mutant animals. The alpha 3 and alpha 7 subunits showed no change in expression among all groups.

Enhanced epileptogenic susceptibility in a genetic model of reactive synaptogenesis: the spastic Han-Wistar rat.

Our laboratory has been studying the spastic Han-Wistar (sHW) rat as a model of neuronal degeneration. Mutant sHW rats display a number of developmental abnormalities that eventually lead to hippocampal pyramidal cell death and synaptic reorganization starting around 30 days of age. The present study examined the contribution of hippocampal reorganization to the expression of seizures induced by systemic injections of kainic acid. Behavioral observations, EEG recordings and hippocampal Fos protein expression in these animals indicated that mutants develop paroxysmal discharges and seizures earlier than controls and the intensity of epileptic manifestations is greater. Kainate injections were lethal in 50% of mutants compared to only 5% of controls. Fos expression was increased approximately twofold in the mutant hippocampus, implicating abnormal excitation in this region. Additional studies in untreated animals indicated that GluR2 mRNA expression was significantly increased throughout the hippocampus in mutant animals, possibly contributing to the enhanced susceptibility to kainate treatment. These results confirm the role of synaptic reorganization in the increased propensity to develop epileptic discharges. Our data also underscore the usefulness of this natural model of cell degeneration and reactive synaptogenesis for understanding the mechanisms of neuronal hyperexcitability.