Presence Of Extracellular Magnesium Research Articles

Extracellular divalent ions modulate TREK-2-like channel conductance in prefrontal pyramidal neurons in rats

Background: The aim of the study was to investigate, with the use of the patch clamp technique, the dependence of the conductance of the TREK-2-like potassium leak channel in the medial prefrontal cortex pyramidal neurons on the presence of extracellular magnesium and calcium ions. It is suspected that TREK-2 channels regulate mood and may be associated with the pathophysiology of depression. Since magnesium and calcium deficiency contribute to depressive symptoms, we investigated how TREK-2- like channel pore propertie change in the absence of divalent cations. Results: Single-channel currents were recorded in a cell-attached configuration in enzymatically dispersed pyramidal neurons of the prefrontal cortex in rats. Spontaneous TREK-2-like channel activity was recorded either in the presence or absence of magnesium and calcium ions in extracellular solution. A significant increase in the inward channel conductance was observed when divalent cations were removed from the extracellular solution. Inward rectification was also increased when the bath temperature was raised to 34-37°C. Conclusions: The study confirmed that the activity of TREK-2-like channels is affected by the presence of magnesium and calcium ions in the extracellular solution. Therefore, in vivo, the TREK-2-like channel may possibly participate in the prefrontal cortex dysfunction associated with the deficiency of divalent cations.

Influence of extracellular magnesium on phagocytosis and free cytosolic Mg levels in differentiated U937 and MH-S cells

The influence of extracellular calcium on phagocytosis is limited, and the involvement of extracellular magnesium is unclear. The role of extracellular calcium and magnesium on phagocytosis efficiency was investigated. Extracellular calcium had no influence on the internalization of 1 μm polystyrene particles by primary monocytes as has been shown before for the human lymphoma-derived, differentiated cell line U937 and murine alveolar macrophage-derived MH-S cells. In contrast, the phagocytosis by differentiated U937 cells was positively affected by the presence of extracellular magnesium whereas that of MH-S cells was not. An extracellular increase in the magnesium level did not cause an increase in the free cytosolic magnesium concentration in either cell line. In contrast to magnesium, extracellular calcium caused an increase in intracellular divalent cation levels in differentiated U937 cells. A phagocytosis-enhancing effect in the extracellular space was observed in relation to extracellular magnesium rather than with an intracellular increase in magnesium levels, indicating that murine and human immune cells might be regulated differently.

Developmental Changes in NMDA-Induced Intrinsic Optical Signals in the Hippocampal Dentate Gyrus of Children with Medically Intractable Seizures

The N-methyl-D-aspartate (NMDA) receptor is one of the ionotropic glutamate receptor subtypes and exhibits a voltage-dependent blockade of its channel function by extracellular magnesium. This magnesium block is known to be absent or weak early in development and is gradually acquired while the brain matures. Interestingly, in adult patients with temporal lobe epilepsy, the magnesium block appears to be altered allowing more current to flow at a negative membrane potential. We are interested whether a similar change might be observed in children’s hippocampi that have frequently been involved in medically intractable seizures. In the present study, we grouped the patients into 2 categories based on the degree of brain maturity: (I) children under the age of 2 (immature, n = 2) and (II) children over the age of 2 (mature, n = 6). Dentate gyri were imaged in real time for intrinsic optical signals with the use of a transmitted light in hippocampal slice preparations. Light transmittance (LT), which reflects a neuronal synaptic depolarization and a concomitant change in cell volume, was calculated. In the immature hippocampus, LT increased significantly in response to NMDA in the presence of extracellular magnesium. However, the mature hippocampi showed little response to NMDA unless magnesium ions were removed from the extracellular artificial cerebrospinal fluid. LT increase was also induced in response to α-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA); however, there were no age-dependent differences in the AMPA induced LT increase. Differential sensitivity of the NMDA receptor to extracellular magnesium between immature and mature hippocampi suggests the probable presence of developmental regulation of magnesium block for the NMDA receptor in the human hippocampus of children with medically intractable seizures.

N-methyl-d-aspartate receptor-mediated calcium accumulation in neocortical neurons

Calcium imaging and patch-clamp recording techniques were used to investigate the relationship between membrane properties and intracellular calcium changes in response to the excitatory amino acid neurotransmitter glutamate. Application of glutamate to cultured neocortical neurons produced concentration-dependent increases in intracellular calcium, membrane depolarization and transmembrane current. At a low concentration (3 microM), glutamate induced only a small depolarization (< 10 mV), yet produced a substantial increase in intracellular calcium. The calcium increase was observed in the presence of extracellular magnesium, was dependent on extracellular calcium, was blocked by an N-methyl-D-aspartate receptor antagonist, and was not affected by manipulation of intracellular calcium stores. This low concentration of glutamate also induced membrane currents that exhibited an N-methyl-D-aspartate-like unconventional voltage dependence. When glutamate was increased to a concentration known to produce excitotoxicity (500 microM), large depolarizations and membrane currents were induced, which rapidly reversed following prolonged glutamate applications. Changes in intracellular calcium in response to 500 microM glutamate had both voltage-sensitive and -insensitive components, and consistently remained elevated following removal of glutamate. These results indicate that low concentrations of glutamate can preferentially activate N-methyl-D-aspartate receptors, leading to increases in intracellular calcium. Functionally this may be involved in N-methyl-D-aspartate receptor responses to ambient extracellular glutamate. In addition, N-methyl-D-aspartate receptor-mediated calcium influx and subsequent depolarization induced by high glutamate concentrations can produce alterations in intracellular calcium homeostasis, which may play an important role in excitotoxicity.

The binaural auditory pathway: excitatory amino acid receptors mediate dual timecourse excitatory postsynaptic currents in the rat medial nucleus of the trapezoid body.

We show here that synaptic transmission to the medial nucleus of the trapezoid body (MNTB) is mediated principally by excitatory amino acid receptors and has two components. A fast excitatory postsynaptic current (EPSC) is mediated by non-NMDA receptors and a slow EPSC is mediated by NMDA receptors. Each neuron receives a large synaptic input (calyx of Held) which produces an EPSC with a mean peak conductance of 37 nS. The somatic location of this synapse gives good resolution of the EPSC timecourse with the fast EPSC decaying with a time constant of 1.1 ms (at 25 degrees C). The slow EPSC exhibits a double exponential decay with time constants of 41 ms and 106 ms and is voltage dependent in the presence of extracellular magnesium. Other smaller EPSCS mediated by NMDA and non-NMDA receptors, and a strychnine-sensitive synaptic current, are also present. Although the intrinsic membrane properties of MNTB neurons (Forsythe & Barnes-Davies (Proc. R. Soc. Lond. B 251, 143 (1993)), preceding paper) promote high-fidelity transmission, we show that voltage-dependent modulation of synaptic transmission can occur. Given the specialization of the calyx of Held, it seems that the NMDA-receptor ion channel complex is not primarily serving to potentiate a subthreshold input, but may be involved in the development and maintenance of this exuberant somatic synapse.

Effect of extracellular Ca2+ and Mg2+ on enzyme release from quiescent fibroblasts during various exposures.

Release of enzymes from human fibroblasts, as a result of either ATP depletion caused by metabolic inhibitors or a direct cell membrane damage, was increased in the absence of extracellular Ca2+. The lack of extracellular Mg2+ resulted only in a small nonsignificant increase of enzyme release during ATP depletion. However, in a Ca2+-free medium the enzyme release was very much enhanced during ATP depletion if Mg2+ was absent concomitantly. The data show that extracellular calcium protects the cells from cell damage, and that the presence of extracellular magnesium in a calcium-free medium protects the cell against energy-dependent cell damage.

Spontaneous Tumor Cytolysis Mediated by Inflammatory Neutrophils: Dependence Upon Divalent Cations and Reduced Oxygen Intermediates

The role of divalent cations and reactive products of the respiratory burst were investigated in spontaneous tumor lysis mediated by inflammatory neutrophils (PMNs). Murine peritoneal PMNs, obtained five hours after intraperitoneal injection of bacteria, conjugated and lysed teratocarcinoma cells in chromium release and single-cell cytotoxicity assays. The presence of extracellular magnesium was required and was sufficient for tumor cell binding to PMNs. Postbinding lytic events depended upon the simultaneous presence of extracellular calcium and magnesium. Catalase and superoxide dismutase inhibited postbinding lytic events, indicating that production of reduced oxygen moieties was important. Scavengers of hydroxyl radicals could inhibit tumor cell binding, but none could affect postbinding lytic events. Neither could inhibitors of myeloperoxidase decrease tumor lysis. The ability of conjugating PMNs to lyse their bound targets correlated with their reduction of nitro blue tetrazolium (NBT). Optimal concentrations of phorbol myristate acetate (PMA) markedly increased the NBT positivity of PMNs and the killing of bound tumor cells. Even with optimal stimulation of the respiratory burst, however, there was still a significant number (19%) of bound targets that escaped lysis, suggesting active resistance to oxygen-mediated tumor cell injury.

Spontaneous tumor cytolysis mediated by inflammatory neutrophils: dependence upon divalent cations and reduced oxygen intermediates

The role of divalent cations and reactive products of the respiratory burst were investigated in spontaneous tumor lysis mediated by inflammatory neutrophils (PMNs). Murine peritoneal PMNs, obtained five hours after intraperitoneal injection of bacteria, conjugated and lysed teratocarcinoma cells in chromium release and single-cell cytotoxicity assays. The presence of extracellular magnesium was required and was sufficient for tumor cell binding to PMNs. Postbinding lytic events depended upon the simultaneous presence of extracellular calcium and magnesium. Catalase and superoxide dismutase inhibited postbinding lytic events, indicating that production of reduced oxygen moieties was important. Scavengers of hydroxyl radicals could inhibit tumor cell binding, but none could affect postbinding lytic events. Neither could inhibitors of myeloperoxidase decrease tumor lysis. The ability of conjugating PMNs to lyse their bound targets correlated with their reduction of nitro blue tetrazolium (NBT). Optimal concentrations of phorbol myristate acetate (PMA) markedly increased the NBT positivity of PMNs and the killing of bound tumor cells. Even with optimal stimulation of the respiratory burst, however, there was still a significant number (19%) of bound targets that escaped lysis, suggesting active resistance to oxygen-mediated tumor cell injury.