Traumatic brain injury results in disparate regions of chondroitin sulfate proteoglycan expression that are temporally limited.

Axonal injury is a major hallmark of traumatic brain injury (TBI), and it seems likely that therapies directed toward enhancing axon repair could potentially improve functional outcomes. One potential target is chondroitin sulfate proteoglycans (CSPGs), which are major axon growth inhibitory molecules that are generally, but not always, up-regulated after central nervous system injury. The current study was designed to determine temporal changes in cerebral cortical mRNA or protein expression levels of CSPGs and to determine their regional localization and cellular association by using immunohistochemistry in a controlled cortical impact model of TBI. The results showed significant increases in versican mRNA at 4 and 14 days after TBI but no change in neurocan, aggrecan, or phosphacan. Semiquantitative Western blot (WB) analysis of cortical CSPG protein expression revealed a significant ipsilateral decrease of all CSPGs at 1 day after TBI. Lower CSPG protein levels were sustained until at least 14 days, after which the levels began to normalize. Immunohistochemistry data confirm previous reports of regional increases in CSPG proteins after CNS injury, seen primarily within the developing glial scar after TBI, but also corroborate the WB data by revealing wide areas of pericontusional tissue that are deficient in both extracellular and perineuronal net-associated CSPGs. Given the evidence that CSPGs are largely inhibitory to axonal growth, we interpret these data to indicate a potential for regional spontaneous plasticity after TBI. If this were the case, the gradual normalization of CSPG proteins over time postinjury would suggest that this may be temporally as well as regionally limited.

Age-dependency of 45calcium accumulation following lateral fluid percussion: acute and delayed patterns.

This study was designed to determine the regional and temporal profile of 45calcium (45Ca2+) accumulation following mild lateral fluid percussion (LFP) injury and how this profile differs when traumatic brain injury occurs early in life. Thirty-six postnatal day (P) 17, thirty-four P28, and 17 adult rats were subjected to a mild (approximately 2.75 atm) LFP or sham injury and processed for 45Ca2+ autoradiography immediately, 6 h, and 1, 2, 4, 7, and 14 days after injury. Optical densities were measured bilaterally within 16 regions of interest. 45Ca2+ accumulation was evident diffusely within the ipsilateral cerebral cortex immediately after injury (18-64% increase) in all ages, returning to sham levels by 2-4 days in P17s, 1 day in P28s, and 4 days in adults. While P17s showed no further 45Ca2+ accumulation, P28 and adult rats showed an additional delayed, focal accumulation in the ipsilateral thalamus beginning 2-4 days postinjury (12-49% increase) and progressing out to 14 days (26-64% increase). Histological analysis of cresyl violet-stained, fresh frozen tissue indicated little evidence of neuronal loss acutely (in all ages), but considerable delayed cell death in the ipsilateral thalamus of the P28 and adult animals. These data suggest that two temporal patterns of 45Ca2+ accumulation exist following LFP: acute, diffuse calcium flux associated with the injury-induced ionic cascade and blood brain barrier breakdown and delayed, focal calcium accumulation associated with secondary cell death. The age-dependency of posttraumatic 45Ca2+ accumulation may be attributed to differential biomechanical consequences of the LFP injury and/or the presence or lack of secondary cell death.

Effects of an N-type calcium channel antagonist (SNX 111; Ziconotide) on calcium-45 accumulation following fluid-percussion injury.

Accumulation of calcium following experimental traumatic brain injury (TBI) has been demonstrated to be a prominent pathophysiological component that can compromise mitochondrial functioning and threaten cell survival. The omega-conopeptide SNX-111, also known as Ziconotide, is a potent antagonist of the voltage-gated N-type calcium channel and has demonstrated significant neuroprotective effects against ischemia-induced neuronal injury. To determine whether this compound would be effective in reducing calcium accumulation associated with TBI, SNX-111 was administered intravenously to rats 1 hour following a moderate (2.2 to 2.75 atm) lateral fluid-percussion injury (or sham) at doses of 1 (n = 30), 3 (n = 31), or 5 (n = 30) mg/kg; another group received 0.9% saline solution (n = 35). Brains were processed for calcium 45 (45Ca) autoradiography at 6, 12, 24, 48, and 96 hours following insult. Optical density measurements of 20 cortical and subcortical regions were analyzed. Injured animals administered saline solution exhibited a significant increase in 45Ca uptake within 12 regions ipsilateral to the site of injury. The most prominent increases were evident throughout the ipsilateral cerebral cortex. SNX-111 reduced the injury-induced calcium accumulation within the ipsilateral cortex in a dose-response fashion when measured at 6, 12, and 48 hours after insult. These drug-induced reductions in calcium accumulation were as high as 75% in the ipsilateral cerebral cortex, and up to 50% in other ipsilateral regions (including thalamus and hippocampus). Consequently, the results suggest that posttraumatic blocking of the voltage-gated N-type calcium channel after injury reduces prolonged, trauma-induced calcium accumulation.

Cerebral metabolism following neonatal or adult hemineodecortication in cats: effect on oxidative capacity using cytochrome oxidase histochemistry.

In order to determine the degree and extent of changes in cerebral oxidative capacity following cerebral hemineodecortication, adult cats which had undergone surgery early postnatally (mean age: 11.4 days) or during adulthood were studied using cytochrome oxidase histochemistry. A total of 18 animals were employed and 50 brain regions were quantified bilaterally using optical densitometry. Although many subcortical regions exhibiting extensive degenerative features revealed lower levels of cytochrome oxidase (C.O.) activity, this reduction was relatively unremarkable compared to intact controls. Nevertheless, it was interesting that this decrease (down to 66-89%) of normal was more pronounced in neonatal-lesioned cats, reaching significance in a number of ipsilateral thalamic nuclei, compared to adult-lesioned animals (91-100% of normal), suggesting a contribution of glial cells to the density of C.O. staining in the latter cats. Regions of the brain spared from degeneration exhibited a bilateral increase in C.O. activity which may reflect the demands for energy to support the anatomical reorganization which is prevalent in these animals. Surprisingly, such increases were more robust within spared regions of the adult-lesioned brain, reaching significance in four ipsilateral and nine contralateral areas with the density of the reaction attaining levels over 125% of control. This may indicate different demands for oxidative metabolism in the adult-lesioned cats. These results enhance our understanding of the mechanism(s) underlying the greater extent of functional sparing or recovery in cats sustaining injury to the cerebral cortex early vs. late in life. In addition, the findings complement our previous companion report on glucose metabolism supporting the concept of energy compartmentalization, which reflects the dynamic interaction between anatomical and functional changes in this age-at-lesion model of recovery.

Effects of traumatic brain injury on the cholinergic system in the rat.

Rats subjected to a mild to moderate fluid percussion injury exhibit memory deficits that are similar to rats that have received lesions of the septohippocampal system. Because the cholinergic system plays a major role in septohippocampal function, we studied the kinetics of the synthetic enzyme for acetylcholine, choline acetyltransferase (ChAT), at 1 h, 24 h, or 5 days after a fluid percussion injury. Decreases in ChAT activity were found in the dorsal hippocampus (25%), frontal (32%), and temporal (23%) cortices 1 h after injury. In the parietal cortex, a greater than 50% increase in ChAT activity was observed at all time intervals assessed. At 5 days after TBI, there was an 18% increase in ChAT activity in the medial septal area. These data provide evidence that a mild to moderate fluid percussion injury produces changes in the cholinergic system in brain areas related to memory.

Cerebral metabolism following neonatal or adult hemineodecortication in cats: I. Effects on glucose metabolism using [14C]2-deoxy-D-glucose autoradiography.

In the cat, cerebral hemispherectomy sustained neonatally results in a remarkable degree of recovery and/or sparing of function as compared with the effects of a similar lesion but sustained in adulthood. We have proposed that this effect is due to a combination of reduced neuronal loss within partially denervated structures and a lesion-induced reorganization of corticofugal projections arising from the remaining intact hemisphere in the neonatally lesioned animal. The current study was designed to assess the physiological consequences of these anatomical changes utilizing [14C]2-deoxy-D-glucose autoradiography. A total of 17 adult cats were studied. Seven animals served as intact controls, five received a left cerebral hemineodecortication as neonates (NH; mean age 11.4 days), and five sustained the same lesion in adulthood (AH). Histological analysis indicated that the lesion was very similar between the two age groups and essentially represented a unilateral hemineodecortication. Local CMRglc (LCMRglc; mumol 100 g-1 min-1) values were calculated for 50 structures bilaterally and indicated that in the remaining intact contralateral (right) cerebral cortex (including all areas measured), AH cats exhibited a significantly (p < 0.05) lower level of LCMRglc (ranging from 20 to 72 mumol 100 g-1 min-1) than NH (ranging from 49 to 81 mumol 100 g-1 min-1). In comparison, the rates of NH cats within the cerebral cortex were very similar to those seen in intact animals (ranging from 48 to 119 mumol 100 g-1 min-1). Ipsilateral to the lesion in AH cats, the structures spared by the resection, including the basal ganglia and thalamus, exhibited LCMRglc rates of between 23 and 69 mumol 100 g-1 min-1, which were significantly lower (p < 0.05) than in NH cats (range 47-72 mumol 100 g-1 min-1). Considering all structures, both age-at-lesion groups exhibited a lower level of metabolism compared with similar measurements for intact control animals (LCMRglc range 45-75 mumol 100 g-1 min-1). However, this depression of glucose metabolism was more pronounced in the AH cats (p < 0.05). These results indicate that following neonatal hemineodecortication, LCMRglc is maintained at a higher level in many regions of the brain than in animals that sustain the same resection in adulthood. This higher level of glucose metabolism in NH animals suggests that the lesion-induced anatomical reorganization of structures not directly injured by the lesion plays a functional role that is probably responsible for the greater degree of recovery and/or sparing of function in these early lesioned cats.

Morphological changes in the thalamus and neocortex of the cat brain after a restricted unilateral fetal neocortical lesion.

In order to study the response of the brain to injury during early development, the neocortex of ten fetal kittens was lesioned at age E43-48, in either the frontal (n = 8) or parieto-occipital (n = 2) areas. The thalamus and neocortex of the lesioned animals were analyzed using quantitative morphometry and compared to intact control cats (n = 10). Ipsilaterally, the volumes of the remaining neocortex and of the thalamus were 26.5% and 25.7% smaller, respectively (P < 0.05). Contralaterally, the neocortex did not change in volume, whereas the thalamus tended to be smaller by a mean of 11.1%. Ipsilaterally, in all four thalamic nuclei studied, the neuronal and glial cell packing densities (NCPD and GCPD) and the cross sectional area of neuronal somata did not differ between lesioned and intact animals except for the principal ventromedial nucleus, where the GCPD was significantly lower (P < 0.05) in lesioned animals. Contralaterally, the NCPD and GCPD did not show any differences between groups, except for the principal ventromedial nucleus, in which the GCPD was lower in lesioned cats (P < 0.05). Furthermore, in the contralateral basal ventromedial nucleus, the cross sectional area of the neuronal somata was smaller in lesioned than in intact animals (P < 0.01). These results indicate loss of neurons and glia in the ipsilateral thalamus and probably in the neocortex. Since, at the time of the cortical resection, transient reciprocal thalamosubplate connections have been established in the cat, the lesion-induced deprivation of subplate target neurons and cortical inputs probably precluded the survival of a substantial number of developing thalamic neurons. In the cortex the hypothetical loss of neurons may, at least partly, be attributed to lesion-induced elimination of target neurons before establishment of corticocortical connections.

Maturation of cerebral oxidative metabolism in the cat: a cytochrome oxidase histochemistry study.

The maturation of brain oxidative capacity was studied in kittens, using cytochrome oxidase histochemistry, at different ages throughout development. Optical densitometry values of reacted tissue were obtained for 50 different structures of the brain. In general, most structures reached adult levels of oxidative capacity by 30 days of age with some motor areas (e.g., cerebellum, red nucleus) exhibiting adult values as early as 7 days of age. Thereafter, some structures (e.g., basal ganglia, thalamus) exhibited levels of cytochrome oxidase activity that exceeded adult values for varying periods of time. These findings indicate regional heterogeneity in the maturation of cerebral oxidative capacity. Furthermore, these maturational patterns appear to correlate well with previous observations from anatomical, physiological and neurobehavioral studies.

Dynamic changes in local cerebral glucose utilization following cerebral conclusion in rats: evidence of a hyper- and subsequent hypometabolic state.

Following cerebral concussion, in which there is no evidence of direct morphological damage, cells are exposed to an increase in extracellular potassium as well as an accumulation of calcium. This concussion-induced ionic flux most likely alters the cellular energy demands thereby modifying metabolic processes. To investigate the metabolic changes after cerebral concussion, local cerebral metabolic rates for glucose (lCMRglc) utilizing [14C]2-deoxy-D-glucose were studied in rats (n = 98; 250-300 g) immediately, 30 min, 6 h, 1, 2, 3, 5 and 10 days following a unilateral frontoparietal fluid percussion (F-P) injury (3.7-4.3 atm). Compared to sham controls, animals exhibited bilateral hypermetabolism immediately following brain injury. However, this effect was more pronounced in structures ipsilateral to the site of F-P and was especially marked for the cerebral cortex (46.6-30.1% higher than control) and hippocampus (90.1-84.4% higher than control). By 30 min post-trauma many ipsilateral regions still showed evidence of hypermetabolism, although their lCMRglc had subsided. Beginning as early as 6 h following injury many regions within the ipsilateral cortex and hippocampus went into a state of metabolic depression (16.4-33.7% of control) which lasted for as long as 5 days. These results indicate that, although not mechanically damaged from the insult, cells exposed to concussive injury dramatically alter their metabolic functioning. This period of post-concussive metabolic dysfunction may delineate a period of time, following injury, during which cells are functionally compromised.

Transplantation of fetal frontal cortex onto degenerating thalamus of cats and kittens.

Tissue from fetal frontal cortex survived after transplantation onto the surface of the left thalamus in 2 kittens and 2 adult cats which 7 days previously had sustained a left cerebral hemispherectomy. There were nerve fiber connections with host tissue (WGA-HRP, Loyez myelin stain) only in the neonatal animals. The grafts contained surviving neurons in all but in one adult cat which survived 301 days. The grafts had little effects on the retrograde ventral thalamic degeneration typically seen following hemispherectomy. However, the dorsal lateral geniculate nucleus adjacent to the transplants showed reduced neuronal loss and gliosis compared to controls. Magnetic resonance imaging was successfully used to visualize the grafts in vivo and suggested a decrease in size as well as changes in composition for a graft systematically followed for 120 days posttransplantation. Cytochrome oxidase histochemistry indicated sustained metabolic activity in transplants containing surviving neurons. This study introduces the cat as a useful model for brain tissue transplantation in a classical, myelinated sensorimotor system.