No abstract

D-serine is a physiological coagonist of N-methyl D-aspartate receptors (NMDARs) that plays a major role in several NMDARdependent events. In this study we investigate mechanisms regulating D-serine production by the enzyme serine racemase (SR). We now report that NMDAR activation promotes translocation of SR to the plasma membrane, which dramatically reduces the enzyme activity. Membrane-bound SR isolated from rat brain is not extracted from the membrane by high detergent and salt concentration, indicating a strong association. Colocalization studies indicate that most membrane-bound SR is located at the plasma membrane and dendrites, with much less SR observed in other types of membrane. NMDAR activation promotes translocation of the cytosolic SR to the membrane, resulting in reduced D-serine synthesis, and this effect is averted by blockade of NMDARs. In primary neuronal cultures, SR translocation to the membrane is blocked by a palmitoylation inhibitor, indicating that membrane binding is mediated by fatty acid acylation of SR. In agreement, we found that SR is acylated in transfected neuroblastoma cells using [ 3 H]palmitate or [ 3 H]octanoic acid as precursors. In contrast to classical S-palmitoylation of cysteines, acylation of SR occurs through the formation of an oxyester bond with serine or threonine residues. In addition, we show that phosphorylation of Thr-227 is also required for steady-state binding of SR to the membrane under basal, nonstimulated condition. We propose that the inhibition of D-serine synthesis caused by translocation of SR to the membrane provides a fail-safe mechanism to prevent NMDAR overactivation in vicinal cells or synapses.glutamate ͉ neurotransmission ͉ octanoylation ͉ palmitoylation ͉ synapse D -serine is a physiological ligand of the coagonist site of NMDARs, mediating several NMDAR-dependent events, including NMDAR neurotransmission (1), neurotoxicity (2, 3), synaptic plasticity (4), and cell migration (5). D-serine is synthesized by serine racemase (SR), an enzyme that directly converts L-into D-serine (6). This enzyme is regulated by interacting proteins, such as the glutamate interacting protein 1 (5), Pick-1 (7), and Golga3 (8), and by nitric oxide produced upon NMDAR activation (9).Despite the many roles attributed to it, the regulation of D-serine signaling is still largely unknown. Furthermore, many questions remain unresolved regarding the distribution of SR and the roles played by glia vs. neurons in D-serine signaling (10). Although the highest levels of endogenous D-serine were shown to be present in brain astrocytes (11), D-serine has also been detected in neurons (2). Recent data using new antibodies against SR (2) and SR knockout mice as negative controls (12) indicate that SR is abundantly expressed in neurons, with highest levels in the cerebral cortex and the hippocampal formation. Moreover, endogenous D-serine released from neuronal cultures lacking significant levels of astrocytes mediates NMDAR-elicited neurotoxicity (2), suggesting that neuron-derived...

Edited by Jesus Avila Keywords:D-Serine NMDA receptor Glutamate Gliotransmission Serine racemase a b s t r a c t Serine racemase (SR) catalyses the synthesis of the transmitter/neuromodulator D-serine, which plays a major role in synaptic plasticity and N-methyl D-aspartate receptor neurotoxicity. We now report that SR is phosphorylated at Thr71 and Thr227 as revealed by mass spectrometric analysis and in vivo phosphorylation assays. Thr71 phosphorylation was observed in the cytosolic and membrane-bound SR while Thr227 phosphorylation was restricted to the membrane fraction. The Thr71 site has a motif for proline-directed kinases and is the main phosphorylation site of SR. Experiments with a phosphorylation-deficient SR mutant indicate that Thr71 phosphorylation increases SR activity, suggesting a novel mechanism for regulating D-serine production.

OBJECTIVE: To evaluate whether removal of a double-balloon device for cervical ripening for 6 compared with 12 hours in women with an unfavorable cervix will result in a shorter time to delivery, similar cervical ripening, and without affecting cesarean delivery rate. METHODS: In a prospective randomized trial, cervical ripening was performed using a double-balloon device. Women were randomized to removal of the device after 6 compared with 12 hours. Primary outcome was time to delivery. Secondary outcomes included mode of delivery, Bishop score, and maternal and neonatal adverse outcomes. A sample size of 100 nulliparous and 100 parous women was required assuming a 95% CI, power of 80%, and mean decrease of 6 hours to delivery between the groups. RESULTS: From March 2017 through February 2019, 688 women were screened, 243 were found eligible, and 197 were randomized as follows: nulliparous cohort (n=101): removal after 6 hours (n=48) compared with removal after 12 hours (n=53); parous cohort (n=96): removal after 6 hours (n=49) compared with removal after 12 hours (n=47). Insertion-to-delivery interval was significantly shorter in the 6-hour group for both nulliparous (25.6±12.8 hours vs 31.4±15.2 hours, P<.04; mean difference 5.8, 95% CI 0.2–11.3), and parous cohorts (18.0±6.8 hours vs 22.6±8.2 hours, P=.003; mean difference 4.7, 95% CI 1.6–7.7). Bishop score change and cesarean delivery rate were similar between groups regardless of parity. The 12-hour group in the combined cohort was associated with higher rates of maternal intrapartum fever (2% vs 10%, P=.02; odds ratio 5.3, 95% CI 1.1–24.8). CONCLUSION: Insertion-to-delivery interval is shorter after 6 compared with 12 hours for both nulliparous and parous women. Cervical ripening with a double-balloon device may be achieved in 6 hours. The longer time was associated with a higher rate of intrapartum fever. Six hours should be considered as standard placement time for double-balloon catheters. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov, NCT03045939.