Abstract
Ketamine causes psychotic episodes and is often used as pharmacological model of psychotic-like behavior in animals. There is increasing evidence that molecular mechanism of its action is more complicated than just N-methyl-D-aspartic acid (NMDA) receptor antagonism and involves interaction with the components of calcium homeostatic machinery, in particular plasma membrane calcium pump (PMCA). Therefore, in this study we aimed to characterize brain region-specific effects of ketamine on PMCA activity, interaction with NMDA receptor through postsynaptic density protein 95 (PSD95) scaffolding proteins and glutamate release from nerve endings. In our study, ketamine induced behavioral changes in healthy male rats consistent with psychotic effects. In the same animals, we were able to demonstrate significant inhibition of plasma membrane calcium ATPase (PMCA) activity in cerebellum, hippocampus and striatum. The expression level and isoform composition of PMCAs were also affected in some of these brain compartments, with possible compensatory effects of PMCA1 substituting for decreased expression of PMCA3. Expression of the PDZ domain-containing scaffold protein PSD95 was induced and its association with PMCA4 was higher in most brain compartments upon ketamine treatment. Moreover, increased PSD95/NMDA receptor direct interaction was also reported, strongly suggesting the formation of multiprotein complexes potentially mediating the effect of ketamine on calcium signaling. We further support this molecular mechanism by showing brain region-specific changes in PSD95/PMCA4 spatial colocalization. We also show that ketamine significantly increases synaptic glutamate release in cortex and striatum (without affecting total tissue glutamate content), inducing the expression of vesicular glutamate transporters and decreasing the expression of membrane glutamate reuptake pump excitatory amino acid transporters 2 (EAAT2). Thus, ketamine-mediated PMCA inhibition, by decreasing total Ca2+ clearing potency, may locally raise cytosolic Ca2+ promoting excessive glutamate release. Regional alterations in glutamate secretion can be further driven by PSD95-mediated spatial recruitment of signaling complexes including glutamate receptors and calcium pumps, representing a novel mechanism of psychogenic action of ketamine.
Highlights
Ketamine, a high affinity non-competitive antagonist of N-methyl-D-aspartic acid receptor (NMDAR) is known to induce schizophrenia-like symptoms (Becker et al, 2003)
The effect of ketamine is mainly attributable to the prefrontal cortex, functional deficits in other brain regions such as hippocampus, cerebellum and striatum are suggested to contribute to drug-induced psychotic symptoms (Lidow, 2003; Javitt, 2010; Sleigh et al, 2014)
We first compared the effect of ketamine treatment on plasma membrane calcium ATPase (PMCA) activity in these brain regions using two independent methods—measurement of ATP hydrolysis and formation of phosphointermediate complex
Summary
A high affinity non-competitive antagonist of N-methyl-D-aspartic acid receptor (NMDAR) is known to induce schizophrenia-like symptoms (Becker et al, 2003). Controlled induction of psychotic-like behavior is widely used to mimic the symptoms and molecular abnormalities of this disease in various animal models (Chatterjee et al, 2012; Koh et al, 2016). Dopaminergic deficits may be secondary to underlying glutamatergic dysfunction, mainly in striatal and prefrontal brain regions (Javitt, 2010; Sleigh et al, 2014), it is more evident that ketamine acts on multiple brain areas and influences the function of components other than NMDAR (Lindefors et al, 1997; Howes and Kapur, 2009; Koh et al, 2016)
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