Signaling mechanisms downstream of quinolinic acid targeting the cytoskeleton of rat striatal neurons and astrocytes

Abstract

The studies of signaling mechanisms involved in the disruption of the cytoskeleton homeostasis were performed in a model of quinolinic acid (QUIN) neurotoxicity in vitro. This investigation focused on the phosphorylation level of intermediate filament (IF) subunits of astrocytes (glial fibrillary acidic protein - GFAP) and neurons (low, medium and high molecular weight neurofilament subunits - NFL, NFM and NFH, respectively). The activity of the phosphorylating system associated with the IFs was investigated in striatal slices of rat exposed to QUIN or treated simultaneously with QUIN plus glutamate receptor antagonists, calcium channel blockers or kinase inhibitors. Results showed that in astrocytes, the action of 100 μM QUIN was mainly due to increased Ca(2+) influx through NMDA and L-type voltage-dependent Ca(2+) channels (L-VDCC). In neuronal cells QUIN acted through metabotropic glutamate receptor (mGluR) activation and influx of Ca(2+) through NMDA receptors and L-VDCC, as well as Ca(2+) release from intracellular stores. These mechanisms then set off a cascade of events including activation of PKA, PKCaMII and PKC, which phosphorylate head domain sites on GFAP and NFL. Also, Cdk5 was activated downstream of mGluR5, phosphorylating the KSP repeats on NFM and NFH. mGluR1 was upstream of phospholipase C (PLC) which, in turn, produced diacylglycerol (DAG) and inositol 3,4,5 triphosphate (IP3). DAG is important to activate PKC and phosphorylate NFL, while IP(3) contributed to Ca(2+) release from internal stores promoting hyperphosphorylation of KSP repeats on the tail domain of NFM and NFH. The present study supports the concept of glutamate and Ca(2+) contribution in excitotoxic neuronal damage provoked by QUIN associated to dysfunction of the cytoskeleton homeostasis and highlights the differential signaling mechanisms elicited in striatal astrocytes and neurons.

Biochemical, histopathological and behavioral alterations caused by intrastriatal administration of quinolic acid to young rats

“…As we have shown previously [45], this effect was mediated by Ca 2+ influx through NMDA channels and by oxidative stress. Additionally, alterations in the homeostasis of the cytoskeleton of astrocytes and neurons were found in rat striatal slices treated with 100 lM QUIN.…”

“…QUIN-treated rats showed decreased corticostriatal [ 3 H]glutamate uptake 30 min after lesion; uptake was restored to sham levels after 1 day in the striatum, but the decrease persisted for 2 weeks in the cerebral cortex. 45 [29]. The spreading of the excitotoxic wave from the zone of lesion to other brain regions suggests spatiotemporal cell damage caused by QUIN for up to 4 weeks after lesion.…”

“…Decreased [ 3 H] glutamate uptake and increased 45 Ca 2+ uptake were detected shortly after injection in the striatum and cerebral cortex. In the hippocampus, increased 45 Ca 2+ uptake preceded the decreased [ 3 H]glutamate uptake, without histopathological alterations. Also, corticostriatal astrogliosis was observed 7 days later, progressing to neuronal death at day 14.…”

“…Behavioral, biochemical and histopathological characteristics of these rats were then assessed at 1, 2, 3 and 4 weeks after surgery. [ 3 H]Glutamate and 45 Ca 2+ uptake were assessed at 30 min, 1, 2, 3 and 4 weeks after surgery. All groups were independent.…”

“…All experiments were performed in triplicate, and the mean was used for the statistical calculations. 45 …”