Protective effect of taurine on the light-induced disruption of isolated frog rod outer segments

Isolated frog rod outer segments (ROS) incubated in a Krebs-bicarbonate medium, and illuminated for 2 h, show a profound alteration in their structure. This is characterized by distention of discs, vesiculation, and a marked swelling. The light-induced ROS disruption requires the presence of bicarbonate and sodium chloride. Replacement of bicarbonate by TRIS or HEPES protects ROS structure. Also, substitution of sodium chloride by sucrose or choline chloride maintains unaltered the ROS structure. Deletion of calcium, magnesium, or phosphate does not modify the effect produced by illumination. An increased accumulation of labeled bicarbonate and tritiated water is observed in illuminated ROS, as compared with controls in the dark. The presence of taurine, GABA, or glycine, at concentrations of 5–25 mM, effectively counteracts the light-induced ROS disruption. Taurine (25 mM) reduces labeled bicarbonate and tritiated water levels to those observed in the dark incubated ROS.     

Regional distribution of cGMP-activated ion channels in the plasma membrane of the rod photoreceptor

In patch-clamp recordings from excised membrane patches, the distribution of cGMP-activated channels in the plasma membrane of the rod photoreceptor was examined. These channels have been shown to be the light-sensitive channels that carry the inward dark current in the rod outer segment, where phototransduction occurs (Matthews, 1987); thus, they are centrally involved in photoreceptor transduction. In the outer segment, cGMP-activated channels were present at high density. In the inner segment, cGMP-activated channels were also present, but their density was much lower than in the outer segment. Calcium-activated potassium channels had the opposite distribution, with a higher density in the inner segment. The results suggest that there is a barrier preventing the unrestricted spread of membrane proteins between the inner and outer segments.     

Separation of potassium and calcium channels in the nerve cell soma membrane]

Calcium inward and potassium outward currents were studied on internally dialysed isolated neurons of the snail Helix pomatia. Different sensitivity of the corresponding channels to changes in external pH was found. This difference was used for separation of their activation regions on the potential axis so that the characteristics of the inward and outward currents could be studied with minimal overlap. It is shown that the outward current channels possess a definite permeability to Tris ions (PTris :PK=0.05). This explains the impossibility to switch off this current by substituting Tris for internal potassium. The channels for the inward calcium current inactivate slowly with a first order kinetic; their instantaneous current-voltage characteristic reveals considerable Goldman-type rectification. The selectivity of the calcium channels to other bivallent cations is Ba:Sr:Ca:Mg=2.8:2.6:1.0:0.2.     

Effects of light and temperature on the response gradient of retinal rod outer segments

Photoreceptor outer segments are continually renewed from base to tip and consequently the membrane at the tip is older than at the base. In parallel with this, there are differences in light responsiveness, and it has been proposed that these may be due to the aforementioned differences in age. We have tested this proposal by manipulating the rates of renewal of the outer segments by exposing Xenopus laevis to different temperature and lighting regimes which are known to affect the rates of their outer segment membrane renewal. We have found that differences of responsiveness are, indeed, correlated with the differences in the age gradients along the outer segments. ©1997 Elsevier Science B.V. All rights reserved.     

The concentration of cytosolic free calcium in vertebrate rod outer segments measured with fura-2

The use of fluorescent indicators such as fura-2 (Grynkiewicz et al., 1985) to measure the cytosolic free calcium activity in retinal rods is complicated by the rods' sensitivity both to the fluorescence and to the light that excites it. By stimulating fluorescence from large numbers of rods in whole, loaded retinas and averaging repeated measurements, however, we have been able to monitor changes in free [Ca2+]i during exposure to nonsaturating lights under physiological conditions. Retinas, isolated from the bullfrog Rana catesbeiana, were loaded with fura-2 by incubation and mounted, receptor-side up, in a perfusion chamber placed on the stage of a specially designed apparatus. A step of light delivered from above, whose wavelength alternated between 340 and 380 nm every 110 msec, excited fluorescence from 24 mm2 of retina and evoked a light response (the aspartate-isolated pIII component of the electroretinogram--ERG). By comparing the fluorescence intensities excited by the 2 wavelengths (corrected for background and dark-noise), the free [Ca2+]i of the rod outer segment was determined. In darkness, the [Ca2+]i of the outer segment was found to be approximately 220 nM. A bright light caused it to fall exponentially to approximately 140 nM, with a time constant of approximately 1.6 sec. The value of [Ca2+]i at the onset of illumination was independent of stimulus intensity over a 2 log-unit range, and in all cases the fall was monotonic. After terminating the illumination, [Ca2+]i rose again to its time-zero value.(ABSTRACT TRUNCATED AT 250 WORDS)     

Electroregulated potassium channels of the somatic membrane of mollusk neuron membranes

The delayed potassium current in mollusc neurons was separated into two components: a noninactivating component and a transient (inactivating) component. A noninactivating potassium current is not affected by changes in temperature, whereas an inactivating component decreases substantially under cooling. Internal Na+ can block the delayed outward potassium current. The blockade of the inactivating potassium current is strongly voltage-dependent. The voltage dependence of the blockade of the noninactivating potassium current is not so clear. These results are consistent with the hypothesis that there are two forms of the delayed potassium current channel. The effect of temperature and internal Na+ on the fast outward current is similar to that on the inactivating component of the delayed potassium current.     

Two types of potassium channels in the PC12 cell line

Activation of K⁺ channels in the PC12 cell line was studied by comparing⁸⁶Rb⁺ efflux under depolarizing and non-depolarizing conditions. Evidence for both Ca²⁺-dependent and voltage-dependent K⁺ channels was obtained by studying depolarization-induced⁸⁶Rb⁺ efflux in solutions of varying Ca²⁺ concentration and in the presence of K⁺ and Ca²⁺ channel blocking agents.     

Protective effect of taurine and zinc on peroxidation-induced damage in photoreceptor outer segments

Exposure of isolated frog rod outer segments (ROS) to ferrous sulfate in a Krebsbicarbonate medium causes a time-dependent disruption of the membrane organization of the discs. Ferrous sulfate also causes ROS swelling and aggregation. Addition of taurine (5–20 mM) and zinc sulfate (250 μM) to the incubation medium markedly protected ROS from the disrupting effect of ferrous sulfate. Of other amino acids tested, only β-alanine had a protective effect on ROS structure. Ferrous sulfate caused an increase in lipid peroxidation, measured by malonaldehyde formation. The protective effect of taurine and zinc is not accompanied by a reduction of lipid peroxidation. Water accumulation occurs as a consequence of the peroxidative action of ferrous sulfate, and this effect was counteracted by taurine and zinc. Ferrous sulfate did not cause damage to ROS structure when incubation was carried out in sucrose-HEPES. Sodium, chloride, and bicarbonate ions caused ferrous sulfate to disrupt ROS structure. It is concluded that taurine and zinc protect ROS membranes from ion and/or water entry occurring as a consequence of membrane lipid peroxidation.     

Evidence for the receptor-operated calcium channels in human platelet plasma membrane

Loading of human platelets with quin2 considerably increases platelet activating factor (PAF)-induced 45Ca2+ uptake. 45Ca2+ binding in the absence of agonists remains unchanged. The data show that PAF stimulates calcium ion influx into platelets, since quin2 provides substantial additional buffer capacity for Ca2+ in the cytoplasm. PAF and prostaglandin endoperoxide H2 analogue, U46619, induce Ba2+ and Mn2+ entry. The entrance of these cations and the rise in [Ca2+]i are both blocked by cAMP elevation and activation of protein kinase C by phorbol ester which indicates a single mechanism for agonist-induced transport of all three cations into platelets. It is assumed that agonists stimulate Ca2+ entry through the receptor-operated channels.     

Separation of the sodium and potassium channels in the surface membrane of mollusk nerve cells]

Characteristics of transmembrane ionic currents under controlled changes in ionic composition of extra-and intracellular medium were studied by means of intracellular dialysis and voltage clamp in isolated neurons from the molluscs Helix pomatia and Limnea stagnalis. The outward potassium currents were eliminated by replacement of intracellular potassium by Tris and the pure inward current could be measured. Replacement of the Ringer solution by NA-free or Ca-free solutions in the extracellular medium made it possible to separate the inward current into additive components, one of which is carried by sodium ions, and the other, by calcium ions. The sodium and calcium inward currents are shown to have different kinetics and potential dependence: taumNa = 1+/-0.5 ms, taumCa = = 3+/-1 ms, tauhNa = 8+/-2 ms, tauhCa = 115+/-10 ms when Vm = 0, GNa = 0.5 when Vm==-21+/-2 mV, GCa = 0.5 when Vm=-8+/-2 mV. Both currents were not altered by tetrodoxin (TTX), however calcium current is specifically blocked by externally applied calcium ions (2 X 10(-3) M), verapamil, D = 600 as well as by fluoride while introduced inside a cell. These data prove the existence of separate systems of sodium and calcium ion-conducting channels in the somatic membrane.     

Development of outer segments and synapses in the rabbit retina

The peripheral retina of rabbits aged 0 to 60 days was studied by electron microscopy. Ribbon and conventional synaptogenesis was studied with serial sections, and the density of synapses of the inner plexiform layer was measured on large (1,500μm²) montages. Photoreceptor and bipolar ribbon synapses seem to develop similarly in that processes of the prospective dyad or triad contact the presynaptic ribbon-containing terminal one at a time. No statistically significant difference in the lengths of ribbon lamellae was found at 11 and 30 days. Conventional synapses appear to result from the aggregation of synaptic vesicles on one side of junctions that first existed as symmetrical membrane densities without vesicles. The length of the synaptic membrane specialization remains constant between 0 and 30 days. The density of inner plexiform layer conventional synapses remains at a low and roughly constant level from 0 to 9 days, after which there is an abrupt increase to a plateau at about 20 days. After nine days the density of ribbon synapses also increases, with an initially steep time course similar to that of conventional synapses. All subcategories of synapse studied (amacrine-to-amacrine, amacrine-to-bipolar, serial, and reciprocal) participate in the general increase between 9 and 20 days. Functional circuits of the inner plexiform layer thus seem to be assembled primarily during the second and third weeks of life.     

Toxins in the characterization of potassium channels

Several recently characterized toxins (apamin, charybdotoxin, dendrotoxin and noxiustoxin) are proving invaluable for establishing what kinds of potassium channel are expressed in neurones, and what the roles of the channels might be.     

Antidepressant-like effect of lamotrigine is reversed by veratrine: a possible role of sodium channels in bipolar depression

Lamotrigine has been found to be efficacious in the acute management of bipolar depression and long-term management of bipolar disorder, especially in delaying depressive recurrence, either as monotherapy or as adjunctive therapy. Lamotrigine is also an antiepileptic drug, and is efficient in the treatment of focal epilepsies. It is thought to act by inhibition of glutamate release through blockade of voltage-sensitivity sodium channels and stabilization of the neuronal membrane. OBJECTIVES: The scope of this study was to determinate if sodium channels are important for lamotrigine and other antidepressant to exert their antidepressant-like function. METHODS: This study assessed the effects of veratrine, a Na(+) channel opener on antidepressant effect of lamotrigine and others antidepressants: two tricyclic antidepressants (TCAs): imipramine, a mixed serotonergic noradrenergic reuptake inhibitor, desipramine, a specific noradrenergic reuptake inhibitor and a SSRI: paroxetine, the most potent selective serotonergic reuptake inhibitor, using an animal model of depression, the forced swimming test. Veratrine (0.125 mg/kg) and lamotrigine (16, 32 mg/kg) or antidepressants (16, 32 mg/kg) were given i.p. 45 and 30 min, respectively, before the test. RESULTS: We observed that when combined with veratrine the antidepressant-like effect of lamotrigine was reversed, but the antidepressant-like effect of the imipramine, desipramine and paroxetine was not changed, indicating that the mechanism of action of lamotrigine is different from that of antidepressants.     

Serum of Isaacs' syndrome suppresses potassium channels in PC-12 cell lines

Blockage of K⁺ channels in nerve terminals by immunoglobulin is the speculated pathomechanism of Isaacs' syndrom. Using patch-clamp technique (whole-cell clamp), we investigated the effects on K⁺ current of serum taken from 2 patients with Isaacs' syndrom employing the clonal cell line PC-12. The addition of a patient's serum to the perfusion solution had little effect on the K⁺ current of P-12 cells. In contrast, K⁺ current was reduced by 25–80% when cells were cultured for 3–6 days with 2% serum as compared to control serum values. Supperession of the K⁺ current appears to develop gradually over the period of culture. Our results suggest that the pathomechanism of Isaacs' syndrome is caused by K⁺ channel suppression via a humoral factor(s) in the serum, which subsequently induces nerve hyperexcitability. © 1996 John Wiley & Sons, Inc.     

Functional characteristics of potassium channels in the somatic membrane of neurons of Lymnaea stagnalis of various ages

It has been found that in old Lymnaea stagnalis neurons the maximal amplitude of the delayed potassium current and the maximal potassium conductance decreases and the kinetics of inactivation of the delayed outward current slows down. It is suggested that these changes may be associated with the age peculiarities of the phospholipid composition of membranes and not related to shifts in the surface area of the nerve cell during aging. The age dependence of the function of potassium channels may underlie the changes in the neuronal function during aging.     

Involvement of ATP-sensitive potassium channels and the opioid system in the anticonvulsive effect of zolpidem in mice

Zolpidem is a hypnotic medication that mainly exerts its function through activating γ-aminobutyric acid (GABA)_A receptors. There is some evidence that zolpidem may have anticonvulsive effects. However, the mechanisms underlying this effect have not been elucidated yet. In the present study, we used the pentylentetrazole (PTZ)-induced generalized seizure model in mice to investigate whether zolpidem can affect seizure threshold. We also further evaluated the roles of ATP-sensitive potassium (K_ATP) channels as well as μ-opioid receptors in the effects of zolpidem on seizure threshold. Our data showed that zolpidem in a dose-dependent manner increased the PTZ-induced seizure threshold. The noneffective (i.e., did not significantly alter the PTZ-induced seizure threshold by itself) doses of K_ATP channel blocker (glibenclamide) and nonselective opioid receptor antagonist (naloxone) were able to inhibit the anticonvulsive effect of zolpidem. Additionally, noneffective doses of either K_ATP channel opener (cromakalim) or nonselective μ-opioid receptor agonist (morphine) in combination with a noneffective dose of zolpidem exerted a significant anticonvulsive effect on PTZ-induced seizures in mice. A combination of noneffective doses of naloxone and glibenclamide, which separately did not affect zolpidem effect on seizure threshold, inhibited the anticonvulsive effects of zolpidem. These results suggest a role for K_ATP channels and the opioid system, alone or in combination, in the anticonvulsive effects of zolpidem.     

Relation between metabolism and the function of ion channels in the nerve cell membrane

The effect of varying intracellular 3.5-cAMP level on steady-state and voltage-dependent transmembrane ionic currents has been studied on both vertebrate and invertebrate nerve cells. A change in 3.5-cAMP level was achieved either by its direct introduction into the cell or by stimulation or inhibition of the activity of different enzymes of the cyclase system. Intracellular cAMP increase was found to activate a steady-state transmembrane current whose early component is related mainly to the increase of sodium and calcium conductance of the membrane, and a late one to increase of potassium conductance (possibly, activated by Ca2+ influx). The decrease of intracellular cAMP concentration (by intracellular dialysis) results in reduction of the potential-activated inward calcium current, whereas its increase restores the current. Restoration of calcium current can be achieved by activation of cellular adenylate cyclase, inhibition of phosphodiesterase or by direct intracellular introduction of the catalytic subunit of cAMP-dependent protein kinase. The evidence is presented that the observed regulatory effects are mediated through cAMP-dependent phosphorylation of proteins of corresponding ionic channels. The increase of intracellular Ca2+ level closely cooperates with this regulatory metabolic system by activation of a number of its enzymatic processes.     

Large‐conductance calcium‐activated potassium channels in purkinje cell plasma membranes are clustered at sites of hypolemmal microdomains

Calcium‐activated potassium channels have been shown to be critically involved in neuronal function, but an elucidation of their detailed roles awaits identification of the microdomains where they are located. This study was undertaken to unravel the precise subcellular distribution of the large‐conductance calcium‐activated potassium channels (called BK, KCa1.1, or Slo1) in the somatodendritic compartment of cerebellar Purkinje cells by means of postembedding immunogold cytochemistry and SDS‐digested freeze‐fracture replica labeling (SDS‐FRL). We found BK channels to be unevenly distributed over the Purkinje cell plasma membrane. At distal dendritic compartments, BK channels were scattered over the plasma membrane of dendritic shafts and spines but absent from postsynaptic densities. At the soma and proximal dendrites, BK channels formed two distinct pools. One pool was scattered over the plasma membrane, whereas the other pool was clustered in plasma membrane domains overlying subsurface cisterns. The labeling density ratio of clustered to scattered channels was about 60:1, established in SDS‐FRL. Subsurface cisterns, also called hypolemmal cisterns, are subcompartments of the endoplasmic reticulum likely representing calciosomes that unload and refill Ca²⁺ independently. Purkinje cell subsurface cisterns are enriched in inositol 1,4,5‐triphosphate receptors that mediate the effects of several neurotransmitters, hormones, and growth factors by releasing Ca²⁺ into the cytosol, generating local Ca²⁺ sparks. Such increases in cytosolic [Ca²⁺] may be sufficient for BK channel activation. Clustered BK channels in the plasma membrane may thus participate in building a functional unit (plasmerosome) with the underlying calciosome that contributes significantly to local signaling in Purkinje cells. J. Comp. Neurol. 515:215–230, 2009. © 2009 Wiley‐Liss, Inc.