endothelin-1 and Movement-Disorders

Neonatal arterial ischemic stroke is one of the more severe birth complications. The injury can result in extensive neurological damage and is robustly associated with later diagnoses of cerebral palsy (CP). An important part of efforts to develop new therapies include the on-going refinement and understanding of animal models that capture relevant clinical features of neonatal brain injury leading to CP. The potent vasoconstrictor peptide, Endothelin-1 (ET-1), has previously been utilised in animal models to reduce local blood flow to levels that mimic ischemic stroke. Our previous work in this area has shown that it is an effective and technically simple approach for modelling ischemic injury at very early neonatal ages, resulting in stable deficits in motor function. Here, we aimed to extend this model to also examine the impact on cognitive function. We show that focal delivery of ET-1 to the cortex of Sprague Dawley rats on postnatal day 0 (P0) resulted in impaired learning in a touchscreen-based test of visual discrimination and correlated with important clinical features of CP including damage to large white matter structures.

Topics: Animals; Animals, Newborn; Association Learning; Atrophy; Brain Ischemia; Cell Count; Cerebral Cortex; Cerebral Palsy; Cognition Disorders; Corpus Striatum; Disease Models, Animal; Endothelin-1; Inflammation; Injections; Microglia; Movement Disorders; Neurons; Perceptual Disorders; Rats; Rats, Sprague-Dawley; Rotarod Performance Test; Vasoconstrictor Agents; White Matter

Recovery of hand function following lesions in the primary motor cortex (M1) is associated with a reorganization of premotor areas in the ipsilesional hemisphere, and this reorganization depends on the size of the lesion. It is not clear how lesion size affects motor representations in the contralesional hemisphere and how the effects in the 2 hemispheres compare. Our goal was to study how lesion size affects motor representations in the ipsilesional and contralesional hemispheres. In rats, we induced lesions of different sizes in the caudal forelimb area (CFA), the equivalent of M1. The effective lesion volume in each animal was quantified histologically. Behavioral recovery was evaluated with the Montoya Staircase task for 28 days after the lesion. Then, the organization of the CFA and the rostral forelimb area (RFA)--the putative premotor area in rats--in the 2 cerebral hemispheres was studied with intracortical microstimulation mapping techniques. The distal forelimb representation in the RFA of both the ipsilesional and contralesional hemispheres was positively correlated with the size of the lesion. In contrast, lesion size had no effect on the contralesional CFA, and there was no relationship between movement representations in the 2 hemispheres. Finally, only the contralesional RFA was negatively correlated with chronic motor deficits of the paretic forelimb. Our data show that lesion size has comparable effects on motor representations in premotor areas of both hemispheres and suggest that the contralesional premotor cortex may play a greater role in the recovery of the paretic forelimb following large lesions.

Topics: Animals; Brain Ischemia; Brain Mapping; Chronic Disease; Disease Models, Animal; Endothelin-1; Forelimb; Functional Laterality; Motor Activity; Motor Cortex; Movement Disorders; Neural Pathways; Neuronal Plasticity; Random Allocation; Rats; Rats, Sprague-Dawley; Recovery of Function

Ethanol is an important risk factor for the occurrence of cerebral ischemia contributing to poor prognosis and inefficacy of drug treatments for stroke-related symptoms. Females have a higher lifetime risk for stroke than males. Moreover, female gender has been associated with increased ethanol consumption during adolescence. In the present study, we investigated whether chronic ethanol exposure during adolescence may potentiate the motor impairments and cortical damage induced by focal ischemia in female rats. We also addressed whether these effects can be mitigated by minocycline, which has been shown to be neuroprotective against different insults in the CNS. Female rats were treated with distilled water or ethanol (6.5 g/kg/day, 22.5% w/v) for 55 days by gavage. Focal ischemia was induced by microinjections of endothelin-1 (ET-1) into the motor cortex. Animals of both groups were treated daily with minocycline (25-50 mg/kg, i.p.) or sterile saline (i.p.) for 5 days, and motor function was assessed using open field, inclined plane and rotarod tests. Chronic ethanol exposure exacerbated locomotor activity and motor coordination impairments induced by focal ischemia in rats. Moreover, histological analysis revealed that microinjections of ET-1 induced pyramidal neuron loss and microglial activation in the motor cortex. Minocycline reversed the observed motor impairments, microglial activation and pyramidal neuron loss in the motor cortex of ischemic rats even in those exposed to ethanol. These results suggest that minocycline induces neuroprotection and functional recovery in ischemic female rats intoxicated with ethanol during adolescence. Furthermore, the mechanism underlying this protective effect may be related to the modulation of neuroinflammation.

Topics: Alcohol-Related Disorders; Animals; Brain Ischemia; Central Nervous System Depressants; Disease Models, Animal; Endothelin-1; Ethanol; Female; Microglia; Minocycline; Motor Activity; Motor Cortex; Movement Disorders; Neurons; Neuroprotective Agents; Pyramidal Cells; Rats, Wistar; Recovery of Function

Despite the availability of numerous transgenic mouse lines to study the role of individual genes in promoting neural repair following stroke, few studies have availed of this technology, primarily due to the lack of a reproducible ischemic injury model in the mouse. Intracortical injections of Endothelin-1 (ET1) a potent vasoconstrictive agent, reliably produces focal infarcts with concomitant behavioral deficits in rats. In contrast, ET1 infarcts in mice are significantly smaller and do not generate consistent behavioral deficits.. We have modified the ET1 ischemia model to target the anterior forelimb motor cortex (aFMC) and show that this generates a reproducible focal ischemic injury in mice with consistent behavioral deficits. Furthermore, we have developed a novel analysis of the cylinder test by quantifying paw-dragging behavior.. ET1 injections which damage deep layer neurons in the aFMC generate reproducible deficits on the staircase test. Cylinder test analysis showed no forelimb asymmetry post-injection; however, we observed a novel paw-dragging behavior in mice which is a positive sign of damage to the FMC.. Previous ET1 studies have demonstrated inconsistent behavioral deficits; however, targeting ET1 injections to the aFMC reliably results in staircase deficits. We show that analysis of paw-dragging behavior in the cylinder test is a more sensitive measure of damage to the FMC than the classical forelimb asymmetry analysis.. We have developed a focal ischemic injury model in the mouse that results in reproducible behavioral deficits and can be used to test future regenerative therapies.

Topics: Animals; Brain Ischemia; Cell Count; Disease Models, Animal; Endothelin-1; Forelimb; Immunohistochemistry; Male; Mice; Motor Cortex; Movement Disorders; Neurons; Random Allocation; Reproducibility of Results; Severity of Illness Index; Stroke

Bradykinesia in upper extremities is associated with a wide variety of motor disorders; however, there are few tasks that assay forelimb movement speed in rodent models. This study describes the bradykinesia assessment task, a novel method to quantitatively measure forelimb speed in rats. Rats were trained to reach out through a narrow slot in the cage and rapidly press a lever twice within a predefined time window to receive a food reward. The task provides measurement of multiple parameters of forelimb function, including inter-press interval, number of presses per trial, and success rate. The bradykinesia assessment task represents a significant advancement in evaluating bradykinesia in rat models because it directly measures forelimb speed. The task is fully automated, so a single experimenter can test multiple animals simultaneously with typically in excess of 300 trials each per day, resulting in high statistical power. Several parameters of the task can be modified to adjust difficulty, which permits application to a broad spectrum of motor dysfunction models. Here we show that two distinct models of brain damage, ischemic lesions of primary motor cortex and hemorrhagic lesions of the dorsolateral striatum, cause impairment in all facets of performance measured by the task. The bradykinesia assessment task provides insight into bradykinesia and motor dysfunction in multiple disease models and may be useful in assessing therapies that aim to improve forelimb function following brain damage.

Topics: Animals; Brain Ischemia; Cerebral Hemorrhage; Conditioning, Operant; Corpus Striatum; Disease Models, Animal; Endothelin-1; Equipment Design; Female; Forelimb; Hypokinesia; Microbial Collagenase; Motor Cortex; Movement Disorders; Psychomotor Performance; Rats; Rats, Sprague-Dawley; Time Factors

Stroke remains one of the most common diseases with a serious impact on quality of life but few effective treatments exist. Mild hypothermia (33°C) is a promising neuroprotective therapy in stroke management. This study investigated whether a delayed short mild hypothermic treatment is still beneficial as neuroprotective strategy in the endothelin-1 (Et-1) rat model for a transient focal cerebral ischemia. Two hours of mild hypothermia (33°C) was induced 20, 60 or 120 minutes after Et-1 infusion. During the experiment the cerebral blood flow (CBF) was measured via Laser Doppler Flowmetry in the striatum, which represents the core of the infarct. Functional outcome and infarct volume were assessed 24 hours after the insult. In this sub-acute phase following stroke induction, the effects of the hypothermic treatment on apoptosis, phagocytosis and astrogliosis were assessed as well. Apoptosis was determined using caspase-3 immunohistochemistry, phagocytic cells were visualized by CD-68 expression and astrogliosis was studied by glial fibrillary acidic protein (GFAP) staining.. Cooling could be postponed up to 1 hour after the onset of the insult without losing its positive effects on neurological deficit and infarct volume. These results correlated with the caspase-3 staining. In contrast, the increased CD-68 expression post-stroke was reduced in the core of the insult with all treatment protocols. Hypothermia also reduced the increased levels of GFAP staining, even when it was delayed up to 2 hours after the insult. The study confirmed that the induction of the hypothermia treatment in the Et-1 model does not affect the CBF.. These data indicate that in the Et-1 rat model, a short mild hypothermic treatment delayed for 1 hour is still neuroprotective and correlates with apoptosis. At the same time, hypothermia also establishes a lasting inhibitory effect on the activation of astrogliosis.

Topics: Animals; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Apoptosis; Brain Infarction; Brain Ischemia; Caspase 3; Cell Count; Cerebrovascular Circulation; Disease Models, Animal; Endothelin-1; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Gliosis; Hypothermia, Induced; Laser-Doppler Flowmetry; Male; Movement Disorders; Neurologic Examination; Random Allocation; Rats; Rats, Wistar; Statistics as Topic; Time Factors

Unilateral sensorimotor cortical (SMC) lesions in rats impair reaching and grasping movements of the contralateral forelimb. These impairments can be improved using motor rehabilitative training on a skilled reaching task, but the training may be far from sufficient to return animals to pre-lesion levels of performance. Because D-amphetamine (AMPH) has been found to promote neuroplastic responses to injury and to be very beneficial when combined with some (but not all) types of rehabilitative training, we asked in this experiment whether it could improve the efficacy of rehabilitative training in skilled reaching. Ten to 14 days after unilateral ischemic (endothelin-1 induced) lesions of the SMC, adult rats were given a 3-week regimen of AMPH (1mg/kg) coupled with daily rehabilitative training on a skilled reaching task, the single pellet retrieval task. AMPH treatment not only dramatically improved reaching performance compared with saline-injected controls, the AMPH treated rats surpassed pre-lesion levels of performance by the end of the rehabilitative training period. The greater performance in AMPH compared to saline-treated rats was still evident at 1 month, but not at 2 and 3 months, after the end of rehabilitative training. Thus, AMPH treatment can greatly enhance the efficacy of rehabilitative training on a skilled reaching task after unilateral SMC lesions, but alternate injection and training regimes may be needed to produce permanent improvements.

Topics: Animals; Brain Ischemia; Central Nervous System Stimulants; Cerebral Infarction; Denervation; Dextroamphetamine; Disease Models, Animal; Dose-Response Relationship, Drug; Endothelin-1; Male; Motor Cortex; Motor Skills; Movement Disorders; Neuronal Plasticity; Paresis; Physical Conditioning, Animal; Rats; Rats, Long-Evans; Recovery of Function; Treatment Outcome

Previous studies have established the usefulness of endothelin-1 (ET-1) for the production of focal cerebral ischemia. The present study assessed the behavioral effects of focal ET-1-induced lesions of the sensorimotor cortex (SMC) in adult rats as well as cellular and structural changes in the contralateral homotopic motor cortex at early (2 days) and later (14 days) post-lesion time points. ET-1 lesions resulted in somatosensory and postural-motor impairments in the contralateral (to the lesion) forelimb as assessed on a battery of sensitive measures of sensorimotor function. The lesions also resulted in the development of a hyper-reliance on the ipsilateral forelimb for postural-support behaviors. In comparison to sham-operated rats, in layer V of the motor cortex opposite the lesions, there were time- and laminar-dependent increases in the surface density of dendritic processes immunoreactive for microtubule-associated protein 2, in the optical density of N-methyl-D-asparate receptor (NMDA) subunit 1 immunoreactivity, and in the numerical density of cells immunolabeled for Fos, the protein product of the immediate early gene c-fos. These findings corroborate and extend previous findings of the effects of electrolytic lesions of the SMC. It is likely that compensatory forelimb behavioral changes and transcallosal degeneration play important roles in these changes in the cortex opposite the lesion, similar to previously reported effects of electrolytic SMC lesions.

Topics: Animals; Brain Ischemia; Corpus Callosum; Endothelin-1; Functional Laterality; Immunohistochemistry; Male; Microtubule-Associated Proteins; Motor Cortex; Movement Disorders; Nerve Degeneration; Neuronal Plasticity; Proto-Oncogene Proteins c-fos; Pyramidal Cells; Rats; Rats, Long-Evans; Receptors, N-Methyl-D-Aspartate; Somatosensory Cortex; Somatosensory Disorders; Time Factors

Tacrolimus (FK506) is a potent immunosuppressant that is presently in clinical use for prevention of allograft rejection. Recently, animal studies reporting significant reductions in the volume of tissue damage associated with cardiac, hepatic, and cerebral ischemia suggest that tacrolimus may also be of use in the clinical management of stroke. In the present study, we examine whether the neuroprotective effects of tacrolimus, as assessed by histological outcome, are accompanied by an amelioration of the skilled motor deficits induced in the rat by middle cerebral artery occlusion (MCAO).. Animals were trained to perform a skilled paw-reaching task before MCAO by perivascular microinjections of endothelin-1. Tacrolimus (1 mg/kg, n = 6) or vehicle (n = 6) was administered by intravenous infusion 1 minute after MCAO. After a 5-day postoperative recovery period, the rats were retested for skilled paw-reaching ability for an additional 9 days.. In vehicle-treated rats, MCAO resulted in a profound bilateral impairment in skilled paw use. Rats treated with tacrolimus, although still impaired, performed significantly better than those treated with vehicle alone (P < .01). Histological analysis, 14 days after occlusion, confirmed the neuroprotective efficacy of tacrolimus with a 66% reduction in the volume of hemispheric brain damage produced by MCAO (P < .01).. The present studies show that tacrolimus not only protects neural tissue from focal cerebral ischemia but also significantly ameliorates the deficits in skilled motor ability produced by this lesion. These data provide further support for the view that tacrolimus may be of use in the treatment of stroke.

Topics: Animals; Brain Ischemia; Cerebral Arteries; Constriction; Endothelin-1; Male; Movement Disorders; Neuroprotective Agents; Psychomotor Performance; Rats; Tacrolimus