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Fluid percussion (FP) brain injury causes spatial memory dysfunction in rats regardless of injury location (midline vs. lateral). Standard histological analysis of the injured brains shows hippocampal neuronal loss after lateral, but not midline FP injury. We have used the optical volume fractionator (OVF) stereological procedure to quantify neuronal loss and glial proliferation within specific subregions of the hippocampus after midline or lateral FP injury. The OVF method is a design-based cell counting procedure, which combines cellular numerical density estimates (from the optical disector) with volume estimates (generated by point counting and the fractionator stereology method) to produce an estimate of the absolute cell number. Fifteen adult male Sprague-Dawley rats were randomly divided into 3 groups (n = 5/group): midline injury, lateral injury and naive. A single fluid percussion pulse was delivered to anesthetized rats in the injured groups. At 14 days post-injury, strict morphological criteria enabled the estimation of neurons, astrocytes, oligodendrocytes, and microglia in defined hippocampal subregions. The results confirm that hippocampal neurons are selectively vulnerable to brain injury, particularly observed as a significant loss in the hilus following both types of injury and in area CA3 after lateral injury. In contrast, the number of astrocytes and oligodendrocytes remains unaffected by brain injury, regardless of subregion. However, the significant increase in microglia number (bilaterally after midline and ipsilateral following lateral injury) suggests that underlying cellular processes continue weeks following injury. The implications of the observed cell population changes are discussed in relation to the reported cognitive deficits associated with both lateral and midline FP brain injury.

Cortical spreading depression (CSD) is associated with release of arachidonic acid, impaired neurovascular coupling, and reduced cerebral blood flow (CBF), caused by cortical vasoconstriction. We tested the hypothesis that the released arachidonic acid is metabolized by the cytochrome P450 enzyme to produce the vasoconstrictor 20-hydroxyeicosatetraenoic acid (20-HETE), and that this mechanism explains cortical vasoconstriction and vascular dysfunction after CSD. CSD was induced in the frontal cortex of rats and the cortical electrical activity and local field potentials recorded by glass microelectrodes, CBF by laser Doppler flowmetry, and tissue oxygen tension (tpO 2 ) using polarographic microelectrodes. 20-HETE synthesis was measured in parallel experiments in cortical brain slices exposed to CSD. We used the specific inhibitor HET0016 (N-hydroxy-NЈ-(4-n-butyl-2-methylphenyl)formamidine) to block 20-HETE synthesis. CSD increased 20-HETE synthesis in brain slices for 120 min, and the time course of the increase in 20-HETE paralleled the reduction in CBF after CSD in vivo. HET0016 blocked the CSD-induced increase in 20-HETE synthesis and ameliorated the persistent reduction in CBF, but not the impaired neurovascular coupling after CSD. These findings suggest that CSD-induced increments in 20-HETE cause the reduction in CBF after CSD and that the attenuation of stimulation-induced CBF responses after CSD has a different mechanism. We suggest that blockade of 20-HETE synthesis may be clinically relevant to ameliorate reduced CBF in patients with migraine and acute brain cortex injuries.

Evoked neural activity correlates strongly with rises in cerebral metabolic rate of oxygen (CMRO 2 ) and cerebral blood flow (CBF). Activity-dependent rises in CMRO 2 fluctuate with ATP turnover due to ion pumping. In vitro studies suggest that increases in cytosolic Ca 2ϩ stimulate oxidative metabolism via mitochondrial signaling, but whether this also occurs in the intact brain is unknown. Here we applied a pharmacological approach to dissect the effects of ionic currents and cytosolic Ca 2ϩ rises of neuronal origin on activitydependent rises in CMRO 2 . We used two-photon microscopy and current source density analysis to study real-time Ca 2ϩ dynamics and transmembrane ionic currents in relation to CMRO 2 in the mouse cerebellar cortex in vivo. We report a direct correlation between CMRO 2 and summed (i.e., the sum of excitatory, negative currents during the whole stimulation period) field EPSCs (ΑfEPSCs) in Purkinje cells (PCs) in response to stimulation of the climbing fiber (CF) pathway. Blocking stimulus-evoked rises in cytosolic Ca 2ϩ in PCs with the P/Q-type channel blocker -agatoxin-IVA (-AGA), or the GABA A receptor agonist muscimol, did not lead to a time-locked reduction in CMRO 2 , and excitatory synaptic or action potential currents. During stimulation, neither -AGA or (-oxo)-bis-(transformatotetramine-ruthenium) (Ru360), a mitochondrial Ca 2ϩ uniporter inhibitor, affected the ratio of CMRO 2 to fEPSCs or evoked local field potentials. However, baseline CBF and CMRO 2 decreased gradually with Ru360. Our data suggest that in vivo activity-dependent rises in CMRO 2 are correlated with synaptic currents and postsynaptic spiking in PCs. Our study did not reveal a unique role of neuronal cytosolic Ca 2ϩ signals in controlling CMRO 2 increases during CF stimulation.