Elizabeth Theriault

MPTP produces reversible disappearance of tyrosine hydroxylase-containing retinal amacrine cells.

To determine whether 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) alters the tyrosine hydroxylase (TH) immunoreactivity of murine dopaminergic retinal amacrine cells, 8-10-week-old C57BL/6J mice were treated with i.p. with saline or cumulative doses of MPTP ranging from 10 to 300 mg/kg. Paraformaldehyde-fixed retinal whole mounts and cross sections were examined using immunochemistry with a tyrosine hydroxylase (TH) or a choline acetyltransferase (ChAT) polyclonal antibody and an avidin-biotin peroxidase reaction. Both TH+ amacrines and ChAT+ retinal neurons showed somal and process morphology and distributions that were commensurate with previous studies of the same or several related species. At 20 days following the MPTP treatment, there was a loss of TH+ amacrines according to a logarithmic relationship relative to MPTP dosage. The loss ranged from 18 to 87% for the dosage range without any decrease in the numbers of ChAT+ neurons. The TH+ amacrines were deleted randomly from the retinas without any peripheral-central predilection. By 273 days after MPTP treatment, the number of TH+ amacrines had returned to values found for age-matched controls demonstrating that the loss of TH immunoreactivity was reversible and occurred without destruction of TH+ amacrines. Computer densitometry revealed that the MPTP-treated TH+ amacrines were divided into two distinct populations: one with normal TH immunodensity levels and a second with TH immunodensity levels below our detection capability. Increasing the MPTP dosage increased the proportion of TH amacrines in the second population. The transient and completely reversible disappearance in the number of TH+ amacrines: (1) appears to form the basis for the decreased concentrations of dopamine and the loss of catecholamine fluorescent neurons previously described for MPTP-treated mouse retinae; (2) may underlie the defects in the electroretinograms of MPTP-treated monkeys, and (3) may result as a response to neurite damage similarly to the alterations in protein synthesis in other central neurons following axonal damage.

Ultrastructural evidence for arteriolar vasospasm after spinal cord trauma.

OBJECTIVE The primary objective of this study was to investigate the potential contribution of vasospasm to the cascade of secondary injury process after traumatic spinal cord injury. Although ischemic factors have been implicated, in that vessel rupture, compression, and intravascular thrombosis are readily identifiable, vasospasm has been more difficult to detect. METHODS The sulcal arterioles in the ventral median fissure of the cervical spinal cord from adult rats were quantitatively examined at the ultrastructural level up to 24 hours after compression injury. RESULTS There were statistically significant changes in the luminal cross-sectional area of sulcal arterioles after spinal cord injury, correlating directly with decreases in length and increases in width of medial smooth muscle cells. A simple mathematical model of postinjury blood flow is presented, suggesting an 80% decrease caused by vasospasm alone. CONCLUSION Our results clearly implicate vasospasm as a contributing factor to secondary injury processes after traumatic spinal cord injury.

Silicone Rubber Microangiography of Acute Spinal Cord Injury in the Rat

The purpose of this study was to investigate the acute changes in the large vessels and microvasculature of the spinal cord after acute clip compression injury in the rat. Nineteen female Wistar rats underwent acute compression of the spinal cord at C8-T1 at 53 g for 1 min. Silicone rubber was injected into the ascending aorta at 15 minutes, 1, 4, or 24 hours after injury. An additional nine rats served as normal controls. The perfused spinal cords were cleared by the alcohol-methylsalicylate technique. The results showed that, in the normal rat, the centrifugal arterial system from the sulcal arteries provided the major blood supply to the gray matter and the lateral and ventral white matter extending all the way to the pial surface. In the normal rat, there were large veins in the posterior columns coursing longitudinally in the parasagittal plane at the base of the posterior columns. The injured spinal cords displayed marked ischemia and hemorrhage at the injury site. The hemorrhage predominated in the gray matter and posterior white columns and extended rostrally and caudally for 2 to 7 mm in each direction from the injury site. Remote hemorrhages originated from damage to the large parasagittal veins in the posterior columns. Extravasations of silicone rubber were frequently seen at the earlier posttraumatic times and often originated from the sulcal arteries or their branches at the injury site. Occluded sulcal arteries were identified at the injury site at 4 and 24 hours.(ABSTRACT TRUNCATED AT 250 WORDS)

Toward an International Initiative for Traumatic Brain Injury Research

The European Commission (EC) and the National Institutes of Health (NIH) jointly sponsored a workshop on October 18-20, 2011 in Brussels to discuss the feasibility and benefits of an international collaboration in the field of traumatic brain injury (TBI) research. The workshop brought together scientists, clinicians, patients, and industry representatives from around the globe as well as funding agencies from the EU, Spain, the United States, and Canada. Sessions tackled both the possible goals and governance of a future initiative and the scientific questions that would most benefit from an integrated international effort: how to optimize data collection and sharing; injury classification; outcome measures; clinical study design; and statistical analysis. There was a clear consensus that increased dialogue and coordination of research at an international level would be beneficial for advancing TBI research, treatment, and care. To this end, the EC, the NIH, and the Canadian Institutes of Health Research expressed interest in developing a framework for an international initiative for TBI Research (InTBIR). The workshop participants recommended that InTBIR initially focus on collecting, standardizing, and sharing clinical TBI data for comparative effectiveness research, which will ultimately result in better management and treatments for TBI.

Number, distribution and neuropeptide content of rat knee joint afferents

Retrograde tracing with Fluoro‐Gold was used to identify the complete population of knee joint afferents in the lumbar dorsal root ganglia of adult female Wistar rats. There was an average of 581±31 (mean± S.D.) afferents supplying each joint. These were found distributed from L1 to L5 with the great majority localised in the L3 and L4 ganglia. Electron microscopy of the posterior articular nerve of the knee revealed an average of 103±15 (mean±S.D.) myelinated and 513±39 unmyelinated axonal profiles. Since about 50–60% of the unmyelinated profiles would be expected to be sympathetic efferents, these numbers are consistent with the numbers of afferents found by Fluoro‐Gold retrograde tracing and suggest that the posterior articular nerve contains about 50% of the total number of knee joint afferents in the rat. Immunohistochemistry revealed that an average of 10% of identified joint afferents expressed substance P‐like immunoreactivity and that 33% expressed calcitonin gene‐related peptide‐like immunoreactivity.

Characterization of axonal ultrastructural pathology following experimental spinal cord compression injury

The present study characterizes axonal pathology associated with traumatic compression injuries of the spinal cord and quantitatively assesses subtypes of axonal pathology in the acute, post-injury period. Eighteen adult female Wistar rats underwent spinal cord compression injury with a 53 g modified aneurysm clip at the C8-T1 segment. Six additional rats served as sham controls. Six experimental animals were sacrificed at each of the three post-injury time points: 15 min, 2 h and 24 h. From all animals, the C8-T1 spinal cord was dissected and processed for both light and electron microscopy. Axonal pathology included periaxonal swelling, organelle accumulation, vesicular myelin, myelin invagination, myelin rupture, and giant axons. Early myelin rupture and the ultrastructural features of giant axons are described here for the first time in the context of spinal cord compression injury. The quantitative analysis characterizes the prevalence of types of axonal pathology over the acute post-injury period and provides evidence for the secondary injury hypothesis regarding the evolution of axonal pathophysiology following trauma.

Effect of nimodipine or methylprednisolone on recovery from acute experimental spinal cord injury in rats

The purpose of the present study was to examine the behavioral, electrophysiologic, and anatomic responses to nimodipine or methylprednisolone treatment of acute experimental spinal cord injury. Four groups of rats were injured at T1 by compressing the cord with a 52-g clip for 1 minute. The treatments were begun 15 minutes after injury, and the animals were observed thereafter for 8 weeks. Nimodipine 0.02 mg/kg/h intravenously (iv) for 8 hours with adjuvant albumen volume expansion, followed by 20 mg/kg nimodipine enterally three times per day for 7 days, produced a moderately better composite score comprising four endpoint parameters than the other treatments which consisted of nimodipine iv for 8 hours only, methylprednisolone 30 mg/kg iv bolus followed by 5.4 mg/kg/h iv for 8 hours, or control.

Confocal imaging of changes in glial calcium dynamics and homeostasis after mechanical injury in rat spinal cord white matter.

Periaxonal glia play an important role in maintaining axonal function in white matter. However, little is known about the changes that occur in glial cells in situ immediately after traumatic injury. We used fluo-3 and confocal microscopy to examine the effects of localized (<0.5 mm) mechanical trauma on intracellular calcium (Ca(i)(2+)) levels in glial cells in a mature rat spinal cord white matter preparation in vitro. At the injury site, the glial Ca(i)(2+) signal increased by 300-400% within 5 min and then irreversibly declined indicating cell lysis and death. In glial cells at sites adjacent to the injury (1.5-2 mm from epicenter), Ca(i)(2+) levels peaked at 10-15 min, and thereafter declined but remained significantly above rest levels. At distal sites (6-9 mm), Ca(i)(2+) levels rose and declined even slower, peaking at 80-90 min. Injury in zero calcium dampened Ca(i)(2+) responses, indicating a role for calcium influx in the generation and propagation of the injury-induced Ca(i)(2+) signal. By 50-80 min post-injury, surviving glial cells demonstrated an enhanced ability to withstand supraphysiological Ca(i)(2+) loads induced by the calcium ionophore A-23187. Glial fibrillary acidic protein (GFAP) and CNPase immunolabeling determined that the glial cells imaged with fluo-3 included both astrocytes and oligodendrocytes. These data provide the first direct evidence that the effects of localized mechanical trauma include a glial calcium signal that can spread along white matter tracts for up to 9 mm within less than 3 h. The results further show that trauma can enhance calcium regulation in surviving glial cells in the acute post-injury period.

Postnatal growth of medial gastrocnemius motoneurons in the kitten

Cat muscle nerves and ventral roots for the hindlimbs show a unimodal distribution of axon diameter at birth, followed, at about 20 days postnatal (dPN), by a marked change to a bimodal distribution resembling that of the adult. However, volumes calculated for motoneuron somata retrogradely labeled with HRP have been reported to be divided into two size populations at birth in the kitten. In the literature it is suggested that a dissociation between axonal and somal growth appears at a very early age. This apparent dissociation, not present in adults, prompted us to examine the somal growth patterns of kitten lumbar motoneurons. In the present report we have examined somal size development in medial gastrocnemius (MG) motor nuclei in 18 cats aged 2 dPN to adulthood using retrogradely transported horseradish peroxidase to label the motoneurons. Measurements of minimum and maximum diameter somal size, volume calculations and a double circle technique relating the diameters of an estimated spherical volume contained within the soma to that of a second spherical volume enclosing the soma clearly distinguish two subpopulations in the adult, a small and a large population. In contrast, in the kitten we show there is a unimodal distribution of small motoneuron somata at birth which at 19-23 dPN differentiates into a bimodal population. This sudden differentiation of somal size coincides with that reported for MG motoneuron axonal calibre, ruling against a neonatal dissociation of somal and axonal size distributions, and appears to correspond to the time of onset of functional characteristics and the histochemical differentiation of fiber types in the MG muscle.

Elevated gene expression in the red nucleus after spinal cord compression injury

Since the mechanism of injury in the majority of human traumatic spinal cord injuries involves rapid cord compression due to bone displacement or fracture-dislocation, we have used a compression injury model to examine the response of adult rat rubrospinal neurons to traumatic spinal cord injury. We have applied in situ hybridization techniques to examine levels of mRNA for cytoskeletal and growth-associated proteins. We report a population of magnocellular red nucleus neurons with elevated levels of T alpha 1 tubulin and GAP-43 up to four weeks post-lesion. In control animals, both probes had very low hybridization signals indistinguishable from background. These and other findings suggest that kinetic compression injuries of the adult spinal cord can result in regeneration-associated gene expression in intrinsic CNS neurons.