P2X4 receptor re-sensitization depends on a protonation/deprotonation cycle mediated by receptor internalization and recycling.

KEY POINTS: Re-sensitization of P2X4 receptors depends on a protonation/de-protonation cycle Protonation and de-protonation of the receptors is achieved by internalization and recycling of P2X4 receptors via acidic compartments Protonation and de-protonation occurs at critical histidine residues within the extracellular loop of P2X4 receptors Re-sensitization is blocked in the presence of the receptor agonist ATP ABSTRACT: P2X4 receptors are members of the P2X receptor family of cation-permeable, ligand-gated ion channels that open in response to the binding of extracellular ATP. P2X4 receptors are implicated in a variety of biological processes, including cardiac function, cell death, pain sensation and immune responses. These physiological functions depend on receptor activation on the cell surface. Receptor activation is followed by receptor desensitization and deactivation upon removal of ATP. Subsequent re-sensitization is required to return the receptor into its resting state. Desensitization and re-sensitization are therefore crucial determinants of P2X receptor signal transduction and responsiveness to ATP. However, the molecular mechanisms controlling desensitization and re-sensitization are not fully understood. In the present study, we provide evidence that internalization and recycling via acidic compartments is essential for P2X4 receptor re-sensitization. Re-sensitization depends on a protonation/de-protonation cycle of critical histidine residues within the extracellular loop of P2X4 receptors that is mediated by receptor internalization and recycling. Interestingly, re-sensitization under acidic conditions is completely revoked by receptor agonist ATP. Our data support the physiological importance of the unique subcellular distribution of P2X4 receptors that is predominantly found within acidic compartments. Based on these findings, we suggest that recycling of P2X4 receptors regulates the cellular responsiveness in the sustained presence of ATP.

Complement C5a Alters the Membrane Potential of Neutrophils during Hemorrhagic Shock.

BACKGROUND: Polymorphonuclear granulocytes (PMN) play a crucial role in host defense. Physiologically, exposure of PMN to the complement activation product C5a results in a protective response against pathogens, whereas in the case of systemic inflammation, excessive C5a substantially impairs neutrophil functions. To further elucidate the inability of PMN to properly respond to C5a, this study investigates the role of the cellular membrane potential of PMN in response to C5a. METHODS: Electrophysiological changes in cellular and mitochondrial membrane potential and intracellular pH of PMN from human healthy volunteers were determined by flow cytometry after exposure to C5a. Furthermore, PMN from male Bretoncelles-Meishan-Willebrand cross-bred pigs before and three hours after severe hemorrhagic shock were analyzed for their electrophysiological response. RESULTS: PMN showed a significant dose- and time-dependent depolarization in response to C5a with a strong response after one minute. The chemotactic peptide fMLP also evoked a significant shift in the membrane potential of PMN. Acidification of the cellular microenvironment significantly enhanced depolarization of PMN. In a clinically relevant model of porcine hemorrhagic shock, the C5a-induced changes in membrane potential of PMN were markedly diminished compared to healthy littermates. Overall, these membrane potential changes may contribute to PMN dysfunction in an inflammatory environment.

State-dependent block of voltage-gated sodium channels by the casein-kinase 1 inhibitor IC261.

Background and Purpose IC261 (3-[(2,4,6-trimethoxyphenyl)methylidenyl]-indolin-2-one) has previously been introduced as an isoform specific inhibitor of casein kinase 1 (CK1) causing cell cycle arrest or cell death of established tumor cell lines. However, it is reasonable to assume that not all antitumor activities of IC261 are mediated by the inhibition of CK1. Meanwhile there is growing evidence that functional voltage-gated sodium channels are also implicated in the progression of tumors as their blockage suppresses tumor migration and invasion of different tumor cell lines. Thus, we asked whether IC261 functionally inhibits voltage-gated sodium channels. Experimental Approach Electrophysiological experiments were performed using the patch-clamp technique at human heart muscle sodium channels heterologously expressed in human TsA cells. Key Results IC261 inhibits sodium channels in a state-dependent manner. IC261 does not interact with the open channel and has only a low affinity for the resting state of the hNav1.5 (human voltage-gated sodium channel; Kr: 120 µM). The efficacy of IC261 strongly increases with membrane depolarisation, indicating that the inactivated state is an important target. The results of different experimental approaches finally revealed an affinity of IC261 to the inactivated state between 1 and 2 µM. Conclusion and Implications IC261 inhibits sodium channels at a similar concentration necessary to reduce CK1δ/ε activity by 50% (IC50 value 1 µM). Thus, inhibition of sodium channels might contribute to the antitumor activity of IC261.

Xenon-induced changes in CNS sensitization to pain.

Electrophysiological investigations of the spinal cord in animals have shown that pain sensitizes the central nervous system via glutamate receptor dependent long-term potentiation (LTP) related to an enhancement of pain perception. To expand these findings, we used functional magnetic resonance (fMRI), blood oxygen level dependent (BOLD) and perfusion imaging in combination with repeated electrical stimulation in humans. Specifically we monitored modulation of somatosensory processing during inhibition of excitatory transmission by ocular application of the glutamate receptor antagonist xenon. BOLD responses upon secondary stimulation increased in mid insular and in primary/secondary sensory cortices under placebo and decreased under xenon treatments. Xenon-induced decreases in regional perfusion were confined to stimulation responsive brain regions and correlated with time courses of xenon concentrations in the cranial blood. Moreover, effects of xenon on behavioral, fMRI and perfusion data scaled with stimulus intensity. The dependence of pain sensitization on sufficient pre-activation reflects a multistage process which is characteristic for glutamate receptor related processes of LTP. This study demonstrates how LTP related processes known from the cellular level can be investigated at the brain systems level.

Autism-associated SHANK3 mutations impair maturation of neuromuscular junctions and striated muscles.

Heterozygous mutations of the gene encoding the postsynaptic protein SHANK3 are associated with syndromic forms of autism spectrum disorders (ASDs). One of the earliest clinical symptoms in SHANK3-associated ASD is neonatal skeletal muscle hypotonia. This symptom can be critical for the early diagnosis of affected children; however, the mechanism mediating hypotonia in ASD is not completely understood. Here, we used a combination of patient-derived human induced pluripotent stem cells (hiPSCs), Shank3Δ11(-/-) mice, and Phelan-McDermid syndrome (PMDS) muscle biopsies from patients of different ages to analyze the role of SHANK3 on motor unit development. Our results suggest that the hypotonia in SHANK3 deficiency might be caused by dysfunctions in all elements of the voluntary motor system: motoneurons, neuromuscular junctions (NMJs), and striated muscles. We found that SHANK3 localizes in Z-discs in the skeletal muscle sarcomere and co-immunoprecipitates with α-ACTININ. SHANK3 deficiency lead to shortened Z-discs and severe impairment of acetylcholine receptor clustering in hiPSC-derived myotubes and in muscle from Shank3Δ11(-/-) mice and patients with PMDS, indicating a crucial role for SHANK3 in the maturation of NMJs and striated muscle. Functional motor defects in Shank3Δ11(-/-) mice could be rescued with the troponin activator Tirasemtiv that sensitizes muscle fibers to calcium. Our observations give insight into the function of SHANK3 besides the central nervous system and imply potential treatment strategies for SHANK3-associated ASD.

In vitro-evaluation of lipid emulsions as vehicles for the administration of xenon: interaction with NMDA receptors.

The noble gas xenon is an antagonist of the NMDA (N-methyl-D-aspartate)-type glutamate receptor which may account for the ideal anesthetic profile and potent neuroprotective properties demonstrated even at subanesthetic concentrations. Because lipid emulsions also promote NMDA antagonistic effects they may serve as ideal carriers for xenon. In this in vitro study, we investigated the efficacy of xenon dissolved in various lipid emulsions (Intralipid, Lipofundin, ClinOleic and Abbolipid on NMDA-evoked currents in cultured cortical neurons. The NMDA receptor blocking property at a clinically relevant concentration seen in the lipid emulsions tested may contribute to the anesthetic, analgetic and neuroprotective effects of xenon administered by way of these lipid carriers. Abbolipid? may serve as the most acceptable carrier since the NMDA antagonistic effect of xenon was enhanced in combination with this emulsion.

Xenon blocks AMPA and NMDA receptor channels by different mechanisms.

The noble gas xenon (Xe) inhibits not only NMDA receptors (NMDARs) but also the two other subtypes of glutamate receptor i.e. AMPA (AMPARs) and kainate receptors. Preliminary studies on AMPARs suggest that Xe sensitivity might be coupled to receptor desensitization. In order to find out if this hypothesis can be applied to all glutamate receptors, we analyzed additional 'non-desensitizing' AMPARs mutants and compared these with homologous mutants of NMDARs. Membrane currents of Neuro2A or SH-SY5Y cells transfected with cDNA encoding AMPA- or NMDA receptors were investigated by whole cell recordings under voltage clamp conditions. Agonists (glutamate, kainate, NMDA) were applied to the cells by means of a rapid perfusion system. Xenon was preincubated for 20 s before testing it in combination with the particular agonist. Xe (3.5 mM) reduced peak and plateau currents of AMPA wild-type receptors [GluR1(i); GluR2(i,Q)] activated for 5 s with 3 mM glutamate, by 45 and 55% respectively. With mutant AMPARs showing greatly diminished or abolished desensitization i.e. GluR1(i)_L497Y, GluR1(i)_A636T(Lc) GluR2(i,Q)_R649E and GluR2(i,Q)_A643T(Lc) the reduction by Xe was significantly smaller and varied by between 4 and 20%. In contrast, no difference in the blocking capacity of Xe was observed comparing wild-type NR1-1a/NR2A receptors with receptors having point mutations within NR2A that substantially slowed (NR2A_A651T(Lc)) or accelerated (NR2A_M823W) receptor desensitization. Thus, our data indicate that in AMPARs channel blockade by Xe is related to desensitization, whereas in NMDARs no evidence for such a relation was found. Thus, Xe seems to exert its inhibiting effect on various ionotropic glutamate receptors by different molecular mechanisms.

Essential role of calcium in vascular endothelial growth factor A-induced signaling: mechanism of the antiangiogenic effect of carboxyamidotriazole.

Vascular endothelial growth factor-A (VEGF-A) plays a major role in tumor angiogenesis and raises the concentration of intracellular free calcium ([Ca2+]i). Carboxyamidotriazole (CAI), an inhibitor of calcium influx and of angiogenesis, is under investigation as a tumoristatic agent. We studied the effect of CAI and the role of [Ca2+]i in VEGF-A signaling in human endothelial cells. VEGF-A induced a biphasic [Ca2+]i signal. VEGF-A increased the level of intracellular inositol 1,4,5-trisphosphate (IP3), which suggests that VEGF-A releases Ca2+ from IP3-sensitive stores and induces store-operated calcium influx. Reduction of either extracellular or intracellular free Ca2+ inhibited VEGF-A-induced proliferation. CAI inhibited IP3 formation, both phases of the calcium signal, nitric oxide (NO) release, and proliferation induced by VEGF-A. CAI prevented neither activation of VEGF receptor-2 (VEGFR-2) (KDR/Flk-1), phospholipase C-g, or mitogen-activated protein kinase (MAP kinase) nor translocation of nuclear factor of activated T cells (NFAT). We conclude that calcium signaling is necessary for VEGF-A-induced proliferation. MAP kinase activation occurs independently of [Ca2+]i but is not sufficient to induce proliferation in the absence of calcium signaling. Inhibition of the VEGF-A-induced [Ca2+]i signal and proliferation by CAI can be explained by inhibition of IP3 formation and may contribute to the antiangiogenic action of CAI. Calcium-dependent NO formation may represent a link between calcium signaling and proliferation.

Open channel block of NMDA receptors by diphenhydramine.

BACKGROUND AND PURPOSE: Diphenhydramine is a well known H1-receptor antagonist that plays a major role in clinical practice. Nowadays, diphenhydramine is primarily applied to prevent nausea but also its sedative and analgesic effects are of clinical importance. As other drugs mediating sedative and analgesic properties partly operate via the inhibition of glutamate receptors, we tested the hypothesis that diphenhydramine, as well interacts with excitatory ionotropic glutamate receptors. EXPERIMENTAL APPROACH: Electrophysiological patch-clamp experiments were performed on glutamate receptors which were heterologously expressed in human TsA cells. KEY RESULTS: Diphenhydramine inhibits NMDA-mediated membrane currents in a reversible and concentration-dependent manner at clinically relevant concentrations. The inhibition occurred in a noncompetitive manner. Diphenhydramine did not compete with NMDA or glycine for their binding sites and half-maximal inhibition was obtained around 25 µM diphenhydramine, independent of the subunit composition. The inhibition was caused by a classical open channel blocking mechanism and varied strongly with the membrane potential. Our results suggest that diphenhydramine most probably interacts with the Mg2+ binding site or a very closely related area of the channel pore. CONCLUSION AND IMPLICATIONS: The data presented here provide evidence that the NMDA receptor antagonism of diphenhydramine contribute to its sedative and potentially LTP-related effects like analgesia and amnesia.

Evaluation of Homobivalent Carbolines as Designed Multiple Ligands for the Treatment of Neurodegenerative Disorders.

Neurodegenerative diseases represent a challenge for biomedical research due to their high prevalence and lack of mechanism-based treatments. Because of the complex pathology of neurodegenerative disorders, multifunctional drugs have been increasingly recognized as potential treatments. We identified homobivalent γ-carbolinium salts as potent inihitors of both cholinesterases, N-methyl-D-aspartate receptors, and monoamine oxidases. Homobivalent γ-carbolines displayed similar structure-activity relationships on all tested targets and may present promising designed multiple ligands for the treatment of neurodegenerative disorders.

Lipid emulsions reduce NMDA-evoked currents.

Membrane currents conducted by the NMDA receptor channels were investigated in cultured cortical neurons and TsA cells transfected with NR1-1a/NR2A subunits of the NMDA receptor. The whole-cell recording technique was used. Current transients evoked by bath application of NMDA for 5 s were characterized by a fast peak and a slow decay to 46.1 +/-15.5% of the peak level at the end. When NMDA was applied in combination with various lipid emulsions (Intralipid, ClinOleic, Lipofundin or Abbolipid, the NMDA-induced currents were reduced, although this reduction did not affect the fast peak, it did affect the decay phase. The amount of reduction depended on the concentration of the lipids (in the case of Abbolipid diluted at 1:40, the current at the end of the 5-s drug application was approximately 2/3 of control). When Abbolipid was applied 40 s before NMDA, peak and late current were reduced to approximately 2/3. The effect of current reduction was the same at either of the two chosen membrane potentials (-80 and +40 mV) which indicates that the effect was not mediated by contamination of the emulsions with Mg(2+). The current reduction produced by Abbolipid was about the same in native neuronal cells and in TsA cells expressing the NR1-1a/NR2A subunits. The current-reducing effect of the lipid emulsions may add to the anesthetic, analgesic and neuroprotective effects seen with hypnotics administered by way of lipid carriers.

Atomoxetine affects transcription/translation of the NMDA receptor and the norepinephrine transporter in the rat brain--an in vivo study.

Attention-deficit/hyperactivity disorder (ADHD) is the most frequently diagnosed neurodevelopmental disorder. The norepinephrine transporter (NET) inhibitor atomoxetine, the first nonstimulant drug licensed for ADHD treatment, also acts as an N-methyl-D-aspartate receptor (NMDAR) antagonist. The compound's effects on gene expression and protein levels of NET and NMDAR subunits (1, 2A, and 2B) are unknown. Therefore, adolescent Sprague Dawley rats were treated with atomoxetine (3 mg/kg, intraperitoneal injection [i.p.]) or saline (0.9%, i.p.) for 21 consecutive days on postnatal days (PND) 21-41. In humans, atomoxetine's earliest clinical therapeutic effects emerge after 2-3 weeks. Material from prefrontal cortex, striatum (STR), mesencephalon (MES), and hippocampus (HC) was analyzed either directly after treatment (PND 42) or 2 months after termination of treatment (PND 101) to assess the compound's long-term effects. In rat brains analyzed immediately after treatment, protein analysis exhibited decreased levels of the NET in HC, and NMDAR subunit 2B in both STR and HC; the transcript levels were unaltered. In rat brains probed 2 months after final atomoxetine exposure, messenger RNA analysis also revealed significantly reduced levels of genes coding for NMDAR subunits in MES and STR. NMDAR protein levels were reduced in STR and HC. Furthermore, the levels of two SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins, synaptophysin and synaptosomal-associated protein 25, were also significantly altered in both treatment groups. This in vivo study detected atomoxetine's effects beyond NET inhibition. Taken together, these data reveal that atomoxetine seems to decrease glutamatergic transmission in a brain region-specific manner. Long-term data show that the compound's impact is not due to an acute pharmacological effect but lasts or even amplifies after a drug-free period of 2 months, leading to altered development of synaptic composition. These alterations might contribute to atomoxetine's clinical effects in the treatment of ADHD, a neurodevelopmental disorder in which synaptic processes and especially a dysregulated glutamatergic metabolism seem to be involved.

Promethazine inhibits NMDA-induced currents - new pharmacological aspects of an old drug.

BACKGROUND AND PURPOSE: The phenothiazine derivative promethazine was first introduced into clinical practice as an antiallergic drug owing to its H1-receptor antagonizing properties. Nowadays, promethazine is primarily used as a sedative and/or as an antiemetic. The spectrum of clinically relevant effects is mediated by different molecular targets. Since glutamate is the predominant excitatory transmitter in the vertebrate brain and involved in alertness control, pain processing, and neurotoxicity we tested the hypothesis that promethazine interacts with excitatory ionotropic glutamate receptors. EXPERIMENTAL APPROACH: Electrophysiological experiments were performed by means of the patch-clamp technique at glutamate receptors heterologously expressed in human TsA cells. KEY RESULTS: Promethazine selectively inhibited NMDA receptors whereas AMPA- and kainate receptors were hardly affected. Inhibition of NMDA-induced membrane currents occurred in a reversible manner with a half-maximal effect at around 20 µM promethazine. The inhibition occurred in a non-competitive manner as it did neither vary with the glutamate nor the glycine concentration. Analysis of the underlying mechanism revealed only a weak dependency on receptor usage, pH value (pH 6.8-7.8), and membrane potential (zδ = 0.44 ± 0.04 according to the Woodhull-model). In line with the latter finding, promethazine did not interact with the Mg(2+) binding site. However, the displacement of promethazine by 9-aminoacridine indicates that promethazine may interact with the channel pore more externally in relation to the Mg(2+) binding site. CONCLUSION AND IMPLICATIONS: Promethazine inhibits NMDA-mediated membrane currents in a reversible and concentration-dependent manner. The results presented here provide evidence that the NMDA receptor antagonism may contribute to clinically relevant effects of promethazine like sedation, analgesia or neuroprotection.

Dose-dependent modulation of apoptotic processes by fluoxetine in maturing neuronal cells: an in vitro study.

OBJECTIVES: Recent studies indicate that the selective serotonin reuptake inhibitor (SSRI) fluoxetine is not solely effective by the instant inhibition of the serotonin transporter (SERT) but also by its influence on mitotic and/or apoptotic processes. METHODS: To investigate the effects of the compound in vitro, we treated neurons from different brain areas with increasing concentrations of fluoxetine. Additionally, human embryonic kidney (HEK-293) cells and HEK-293 cells stably expressing the SERT were used. Cell viability was quantified by MTT-assay and apoptosis via fluorescence-activated cell-sorting analyses. Fluoxetine's effect on the γ-aminobutyric acid (GABA) receptor was electrophysiologically investigated to test the hypothesis if a GABA-mimetic effect exists that might lead - additionally to the well-known N-methyl-D-aspartate (NMDA)-antagonism - to increased apoptosis in immature neurons. RESULTS: In hippocampal, cortical, and both types of HEK-293 cells, viability decreased and apoptosis increased in a dose-dependent manner (0.5-75 µM). In contrast, in mesencephalic and striatal cells the viability was unchanged or even slightly stimulated up to 20 µM fluoxetine. An anti-apoptotic effect of concentrations below 10 µM was observed in these cells. The GABA(A) receptor was directly activated by fluoxetine. CONCLUSIONS: We conclude that fluoxetine affects apoptotic processes independently from SERT expression. Since especially the combined GABA-mimetic and NMDA-antagonistic effects increase apoptosis in developing neuronal cells, whereas both effects are neuroprotective in adult neurons we hypothesise that these mechanisms explain the discrepancy of in vitro and in vivo studies.

Atomoxetine acts as an NMDA receptor blocker in clinically relevant concentrations.

BACKGROUND AND PURPOSE: There is increasing evidence that not only the monoaminergic but also the glutamatergic system is involved in the pathophysiology of attention-deficit hyperactivity disorder (ADHD). Hyperactivity of glutamate metabolism might be causally related to a hypoactive state in the dopaminergic system. Atomoxetine, a selective noradrenaline reuptake inhibitor, is the first non-stimulant approved for the treatment of this disorder. Here we have evaluated the effects of atomoxetine on glutamate receptors in vitro. EXPERIMENTAL APPROACH: The whole-cell configuration of the patch-clamp technique was used to analyse the effect of atomoxetine on N-methyl-d-aspartate (NMDA) receptors in cultured rodent cortical and hippocampal neurons as well as on NMDA receptors heterologously expressed in human TsA cells. KEY RESULTS: Atomoxetine blocked NMDA-induced membrane currents. Half-maximal inhibition emerged at about 3 microM which is in the range of clinically relevant concentrations found in plasma of patients treated with this drug. The inhibition was voltage-dependent, indicating an open-channel blocking mechanism. Furthermore, the inhibitory potency of atomoxetine did not vary when measured on NMDA receptors from different brain regions or with different subunit compositions. CONCLUSIONS AND IMPLICATIONS: The effective NMDA receptor antagonism by atomoxetine at low micromolar concentrations may be relevant to its clinical effects in the treatment of ADHD. Our data provide further evidence that altered glutamatergic transmission might play a role in ADHD pathophysiology.

Evidence that Xenon does not produce open channel blockade of the NMDA receptor.

Previous studies had not excluded the possibility that the mechanism by which Xenon (Xe) blocks N-methyl-D-aspartate (NMDA) receptors might be that of an open-channel blocker. We tested this possibility on mutant NMDA receptors carrying an alanine (A) to cysteine (C) mutation located within the SYTANLAAF-motif of the third transmembrane region (TM3). This mutation was shown to yield constitutively open ion channels after modification with a thiol-modifying reagent. We expressed such mutant channels in Neuro2A cells and recorded glutamate (50 microM)-induced currents in the whole cell recording mode. Although Xe (3.5 mM) blocked the currents through the wild-type receptor NR1-1a/NR2A and NR1-1a/NR2B by approximately 40% and those through the mutant receptors NR1-1a/NR2A(A650C) or NR1-1a/NR2B(A651C) by approximately 30%, it was unable to block the currents through the methane thiosulfonate etyhlammonium-modified mutant receptors. On the other hand, established open-channel blockers of the NMDA receptor such as MK-801 (1 microM) or Mg ions (Mg(2+); 1 mM) were able to block these permanently open channels. These results suggest that Xe does not act as a classical open-channel blocker at the NMDA receptor.

Rapid increase of glial glutamate uptake via blockade of the protein kinase A pathway.

Glutamate is the main excitatory neurotransmitter in the vertebrate central nervous system. Removal of this transmitter from the synaptic cleft by glial and neuronal transporter systems plays an important role in terminating glutamatergic neurotransmission. The effects of different activators and blockers of PKA and PKC on glutamate uptake were studied in primary glial cells cultivated from the rat cortex using the patch-clamp recording technique and immunocytochemical methods. GF 109203X enhances glutamate-induced membrane currents in a concentration- and time-dependent manner. After pre-application for 40 s the maximal transport capacity was increased by 30-80%. The estimated Km-value of the transport system did not change after drug application and the enhanced glutamate uptake was reversible within a few minutes upon washout. Activators and blockers of the PKC pathway did not affect glutamate uptake, whereas H89, a selective blocker of PKA, mimicked the effects of GF 109203X, indicating involvement of the protein kinase A pathway. The GF 109203X-induced increase in transport capacity is likely to be mediated by GLAST since the GLT-1 selective blocker dihydrokainate was unable to block basal or stimulated glutamate uptake. Furthermore, the increase in transport activity may well be based on an increase in cell surface expression of the transporter protein since preincubation with cytochalasin-B, a protein that blocks actin polymerization, almost completely abolished the effect of GF 109203X and H89. These results indicate that GF 109203X and H89 enhance glial glutamate uptake via blockade of the PKA. The described effect may affect glutamatergic neurotransmission by reducing the glutamate concentration in the synaptic cleft.

Activation of neuronal N-methyl-D-aspartate receptor channels by lipid emulsions.

UNLABELLED: Lipid emulsions are widely used as carriers for hypnotics such as propofol, etomidate, and diazepam. It is assumed that the emulsions alone exert no effect on cellular functions nor influence the pharmacokinetics, pharmacodynamics, or anesthetic and analgetic potency of the hypnotics they carry. To elucidate possible interactions between lipid emulsions and cell membranes, in particular membrane-bound proteins, we investigated the effects of commercially available lipid emulsions on the cell membranes of cultured cortical neurons from the mouse by using the whole-cell configuration of the patch-clamp technique. Of nine lipid emulsions tested, three, i.e., Intralipid, Structolipid, and, to a much lesser extent, Abbolipid, activated membrane currents in the neuronal cells in a dilution-dependent manner. The emulsion-induced currents were not affected by picrotoxin or bicuculline but were inhibited by DL-AP5 and ketamine. The voltage dependence of the currents was influenced by the presence of Mg(2+) in a way that is typical for currents conducted by N-methyl-D-aspartate receptor channels. We conclude that Intralipid, Structolipid, and Abbolipid activate N-methyl-D-aspartate receptor channels in cortical neurons. IMPLICATIONS: Lipid emulsions are widely used as carriers for hypnotics such as propofol, etomidate, or diazepam. We tested nine commercially available lipid emulsions and demonstrate that three of them--Intralipid, Structolipid, and Abbolipid--activate NMDA receptor channels in the membranes of cortical neuronal cells.