No abstract

These results suggest that currently hypothesized mechanisms of memantine and ketamine action should be reconsidered and that NR1/2C and/or NR1/2D receptors play a more important role in cortical physiology and pathology than previously appreciated.

Although many nervous system disorders are associated with N -methyl-d-aspartate (NMDA) receptor overactivation, pharmacological inhibition of NMDA receptors has typically demonstrated limited clinical value due to debilitating psychotomimetic side-effects. Memantine, however, induces far fewer behavioural side-effects than other NMDA receptor channel blockers such as ketamine, and slows the progressive cognitive decline associated with Alzheimer's disease. Memantine and ketamine inhibit NMDA receptors with similar affinity and kinetics. A prominent mechanistic difference between memantine and ketamine is the degree to which they are 'trapped' within the closed channel of NMDA receptors following removal of agonist: ketamine becomes trapped in nearly all NMDA receptors to which it was bound before agonist removal, whereas some bound memantine molecules dissociate after agonist removal, a phenomenon called partial trapping. Here we investigated the mechanism underlying partial trapping of memantine by recombinant NR1/2A NMDA receptors. We found that memantine dissociation from NR1/2A receptors after agonist removal (the process that results in partial trapping) followed an exponential time course with τ = 0.79 ± 0.32 s. Neither membrane voltage depolarization nor maintained presence of memantine after agonist removal affected partial trapping, suggesting that partial trapping does not result from memantine escape through open channels. We tested the hypothesis that partial trapping results from binding of memantine to two sites, a superficial 'non-trapping' site and a deep 'trapping' site, which cannot be occupied simultaneously. This hypothesis was supported by the lack of ketamine binding to the superficial site, the voltage dependence of partial trapping, and the effect on partial trapping of a mutation near the deep site. The superficial binding site for memantine may, by causing partial trapping, contribute to memantine's unique therapeutic utility.

Extremely mild hypothermia to 36.0°C is not thought to appreciably differ clinically from 37.0°C. However, it is possible that 36.0°C stimulates highly sensitive hypothermic signaling mechanism(s) and alters biochemistry. To the best of our knowledge, no such ultra-sensitive pathway/mechanisms have been described. Here we show that cold stress protein RNA Binding motif 3 (RBM3) increases in neuron and astrocyte cultures maintained at 33°C or 36°C for 24 or 48h, compared to 37°C controls. Neurons cultured at 36°C also had increased global protein synthesis (GPS). Finally, we found that melatonin or fibroblast growth factor 21 (FGF21) augmented RBM3 upregulation in young neurons cooled to 36°C. Our results show that a 1°C reduction in temperature can induce pleiotropic biochemical changes by upregulating GPS in neurons which may be mediated by RBM3 and that this process can be pharmacologically mimicked and enhanced with melatonin or FGF21.

Neuroprotective cold-shock proteins (CSPs) are abundant in the normothermic neonatal rodent brain but decrease with advancing neurodevelopmental age and are low or absent in the adult brain. It has not been established if neurodevelopmental age alters the baseline expression of CSPs in the human brain. Here, we tested the hypothesis that protein levels of RNA-binding motif 3 (RBM3), reticulon-3 (RTN3), and cold-induced RNA-binding protein (CIRBP) are abundant in the normothermic developing human brain but low-to-absent in adults. We also tested if β-klotho (KLB) is expressed in the developing brain; KLB functions as a coreceptor that controls tissue-specific binding and activity of the systemically circulating thermogenic hormone fibroblast growth factor 21 (FGF21), and is predominantly expressed in the liver, pancreas, and in adipose cells. Methods: Hippocampi and anterior prefrontal cortices (aPFCs/BA10) from a total of 20 male and 20 female subjects were obtained from the NIH NeuroBioBank. CSP and KLB levels were measured in: infants < 1 year old (n = 8), toddlers aged 1–2 years (n = 8), children aged 3–5 years (n = 7), 18-year-old adolescents (n = 8), and adults aged 31–34 years (n = 8). An equal number of male and female (n = 4 each) samples were pooled into each age group, except in the 3- to 5-year-olds which comprised 3 male and 4 female specimens due to sample availability. In total, 78 whole-brain tissues were dissociated using a bead-based Precellys homogenizer to generate equivalent homogenates, and levels of protein targets subsequently analyzed by Western blotting. Results: Infants had the highest levels of RBM3 and other CSPs in the brain compared to all other ages. In the hippocampus, CSPs were detected predominantly in infants. In the aPFC, CSP levels were highest in infants, moderate-to-low in toddlers/children, and below assay detection limits in adolescents/adults. Germane to the thermogenic FGF21/KLB signaling axis, our results confirm that KLB is absent in the adult hippocampus/aPFC as reported by others. In contrast, we report for the first time that KLB is abundant in the early developing human brain; KLB levels were highest in the infant hippocampus/aPFC and moderately expressed in toddlers. RBM3 is a potent neuroprotective CSP. Thus, the impact of these findings on the observed efficacy of therapeutic hypothermia in neonatal brain injury merits further investigation.