Lithium distribution across the membrane of motoneurons in the isolated frog spinal cord

Lithium sensitive microelectrodes were used to investigate the transmembrane distribution of lithium ions (Li+) in motoneurons of the isolated frog spinal cord. After addition of 5 mmol·l–1 LiCl to the bathing solution the extracellular diffusion of Li+ was measured. At a depth of 500 m, about 60 min elapsed before the extracellular Li+ concentration approached that of the bathing solution. Intracellular measurements revealed that Li+ started to enter the cells soon after reaching the motoneuron pool and after up to 120 min superfusion, an intra — to extracellular concentration ratio of about 0.7 was obtained. The resting membrane potential and height of antidromically evoked action potentials were not altered by 5 mmol·l–1 Li+

Inhibitory postsynaptic actions of taurine, GABA and other amino acids on motoneurons of the isolated frog spinal cord

The actions of glycine, GABA, α-alanine, β-alanine and taurine were studied by intracellular recordings from lumbar motoneurons of the isolated spinal cord of the frog. All amino acids tested produced a reduction in the amplitude of postsynaptic potentials, a blockade of the antidromic action potential and an increase of membrane conductance. Furthermore, membrane polarizations occurred, which were always in the same direction as the IPSP. All these effects indicate a postsynaptic inhibitory action of these amino acids. When the relative strength of different amino acids was compared, taurine had the strongest inhibitory potency, followed by β-alanine, α α-alanine, GABA and glycine. Topically applied strychnine and picrotoxin induced different changes of postsynaptic potentials, indicating that distinct inhibitory systems might be influenced by these two convulsants. Interactions with amino acids showed that picrotoxin selectively diminished the postsynaptic actions of GABA, while strychnine reduced the effects of taurine, glycine, α- and β-alanine. But differences in the susceptibility of these amino acid actions to strychnine could be detected: the action of taurine was more sensitively blocked by strychnine compared with glycine, α- and β-alanine. With regard to these results the importance of taurine and GABA as transmitters of postsynaptic inhibition on motoneurons in the spinal cord of the frog is discussed

Effects of guanidine on synaptic transmission in the spinal cord of the frog

The effects of guanidine on motoneurons of the isolated frog spinal cord were studied by adding the drug to the solution bathing the cord during intracellular recording. Guanidine (5·10–4 M) did not alter the membrane potential of motoneurons. The main effect was a marked increase of the amplitudes and frequencies of small spontaneously occurring inhibitory postsynaptic potentials. The hyperpolarizing component of postsynaptic potentials evoked by stimulation of dorsal roots was also enhanced by guanidine. Higher concentrations of guanidine (5·10–3 M) resulted in a very large and irreversible increase of the small spontaneously occurring inhibitory potentials, which now appeared in a regular, rhythmic pattern. The effects of guanidine could easily be blocked by increasing the magnesium ions (15 mM) in the bath solution. These results indicate that guanidine facilitates the release of an inhibitory transmitter in afferent terminals of the frog spinal cord either by a direct action on these terminals or indirectly by an action on nerve endings impinging on inhibitory interneurons

An improved method for constructing and selectively silanizing double-barreled, neutral liquid-carrier, ion-selective microelectrodes

We describe an improved, efficient and reliable method for the vapour-phase silanization of multi-barreled, ion-selective microelectrodes of which the silanized barrel(s) are to be filled with neutral liquid ion-exchanger (LIX). The technique employs a metal manifold to exclusively and simultaneously deliver dimethyldichlorosilane to only the ion-selective barrels of several multi-barreled microelectrodes. Compared to previously published methods the technique requires fewer procedural steps, less handling of individual microelectrodes, improved reproducibility of silanization of the selected microelectrode barrels and employs standard borosilicate tubing rather than the less-conventional theta-type glass. The electrodes remain stable for up to 3 weeks after the silanization procedure. The efficacy of a double-barreled electrode containing a proton ionophore in the ion-selective barrel is demonstrated in situ in the leaf apoplasm of pea (Pisum) and sunflower (Helianthus). Individual leaves were penetrated to depth of ~150 μm through the abaxial surface. Microelectrode readings remained stable after multiple impalements without the need for a stabilizing PVC matrix

Oxygen-glucose deprivation induces ATP release via maxi-anion channels in astrocytes

ATP represents a major gliotransmitter that serves as a signaling molecule for the cross talk between glial and neuronal cells. ATP has been shown to be released by astrocytes in response to a number of stimuli under nonischemic conditions. In this study, using a luciferin-luciferase assay, we found that mouse astrocytes in primary culture also exhibit massive release of ATP in response to ischemic stress mimicked by oxygen-glucose deprivation (OGD). Using a biosensor technique, the local ATP concentration at the surface of single astrocytes was found to increase to around 4 μM. The OGD-induced ATP release was inhibited by Gd3+ and arachidonic acid but not by blockers of volume-sensitive outwardly rectifying Cl− channels, cystic fibrosis transmembrane conductance regulator (CFTR), multidrug resistance-related protein (MRP), connexin or pannexin hemichannels, P2X7 receptors, and exocytotic vesicular transport. In cell-attached patches on single astrocytes, OGD caused activation of maxi-anion channels that were sensitive to Gd3+ and arachidonic acid. The channel was found to be permeable to ATP4− with a permeability ratio of PATP/PCl = 0.11. Thus, it is concluded that ischemic stress induces ATP release from astrocytes and that the maxi-anion channel may serve as a major ATP-releasing pathway under ischemic conditions

ATP release via anion channels

ATP serves not only as an energy source for all cell types but as an ‘extracellular messenger-for autocrine and paracrine signalling. It is released from the cell via several different purinergic signal efflux pathways. ATP and its Mg2+ and/or H+ salts exist in anionic forms at physiological pH and may exit cells via some anion channel if the pore physically permits this. In this review we survey experimental data providing evidence for and against the release of ATP through anion channels. CFTR has long been considered a probable pathway for ATP release in airway epithelium and other types of cells expressing this protein, although non-CFTR ATP currents have also been observed. Volume-sensitive outwardly rectifying (VSOR) chloride channels are found in virtually all cell types and can physically accommodate or even permeate ATP4- in certain experimental conditions. However, pharmacological studies are controversial and argue against the actual involvement of the VSOR channel in significant release of ATP. A large-conductance anion channel whose open probability exhibits a bell-shaped voltage dependence is also ubiquitously expressed and represents a putative pathway for ATP release. This channel, called a maxi-anion channel, has a wide nanoscopic pore suitable for nucleotide transport and possesses an ATP-binding site in the middle of the pore lumen to facilitate the passage of the nucleotide. The maxi-anion channel conducts ATP and displays a pharmacological profile similar to that of ATP release in response to osmotic, ischemic, hypoxic and salt stresses. The relation of some other channels and transporters to the regulated release of ATP is also discussed