Spatial distribution of spermine/spermidine content and K(+)-current rectification in frog retinal glial (Müller) cells.

CMBN, Department of Biochemistry, School of Medicine, Universidad Central del Caribe, Bayamon, Puerto Rico.
Glia (Impact Factor: 6.03). 08/2000; 31(1):84-90. DOI: 10.1002/(SICI)1098-1136(200007)31:13.0.CO;2-7
Source: PubMed


Previous studies in retinal glial (Müller) cells have suggested that (1) the dominant membrane currents are mediated by K(+) inward-rectifier (Kir) channels (Newman and Reichenbach, Trends Neurosci 19:307-312, 1996), and (2) rectification of these Kir channels is due largely to a block of outward currents by endogenous polyamines such as spermine/spermidine (SPM/SPD) (Lopatin et al., Nature 372:366-369, 1994). In frog Müller cells, the degree of rectification of Kir-mediated currents is significantly higher in the endfoot than in the somatic membrane (Skatchkov et al., Glia 27:171-181, 1999). This article shows that in these cells there is a topographical correlation between the local cytoplasmic SPM/SPD immunoreactivity and the ratio of inward to outward K(+) currents through the surrounding membrane area. Throughout the retina, Müller cell endfeet display a high SPM/SPD immunolabel (assessed by densitometry) and a large inward rectification of K(+) currents, as measured by the ratio of inward to outward current produced by step changes in [K(+)](o). In the retinal periphery, Müller cell somata are characterized by roughly one-half of the SPM/SPD immunoreactivity and K(+)-current rectification as the corresponding endfeet. In the retinal center, Müller cell somata are virtually devoid of both SPM/SPD immunolabel and K(+)-current inward rectification. Comparing one region of the retina with another, we find an exponential correlation between the local K(+) rectification and the local SPM/SPD content. This finding suggests that the degree of inward rectification in a given membrane area is determined by the local cytoplasmic polyamine concentration.

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    • "In the central nervous system, spermine/spermidine is predominantly localized in astrocytes (Laube and Veh 1997). Studies from retinal Müller glia have shown that these endogenous polyamines block the outward rectification of K ir 4.1 and also modify gap junction coupling of hippocampal astrocytes in situ (Benedikt et al. 2012; Skatchkov et al. 2000). Additionally , an additive inhibition of Kir4.1 and TASK-1 two-pore domain K ϩ channel could be achieved by their respective channel blockers in Müller glia (Kucheryavykh et al. 2008). "
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    ABSTRACT: Mature astrocytes exhibit a linear current-to-voltage (I-V) relationship K(+) membrane conductance (passive conductance) and an extremely low membrane resistance (RM) in situ. The combination of these electrophysiological characteristics establishes a highly negative and stable membrane potential that is essential for the basic functions, such as K(+) spatial buffering and neurotransmitter uptake. However, astrocytes are coupled extensively in situ. It remains to be determined whether the observed passive behavior and low RM are attributable to the intrinsic properties of membrane ion channels or to gap junction coupling in functionally mature astrocytes. In the present study, freshly dissociated hippocampal tissues were used as a new model to examine this basic question in young adult animals. The morphologically intact single astrocytes could be reliably dissociated from P21 and older animals. At this animal age, dissociated single astrocytes exhibit a similar passive conductance and resting membrane potential as astrocytes do in situ. To precisely measure the RM from single astrocytes, dual patch single astrocyte recording was performed. We show that dissociated single astrocytes exhibit a similarly low RM as syncytial coupled astrocytes. Functionally, the symmetric expression of high K(+) conductance enabled rapid change in the intracellular K(+) concentrations in response to changing K(+) driving force. Altogether, we demonstrate that freshly dissociated tissue preparation is a highly useful model for study of the functional expression and regulation of ion channels, receptors, and transporters in astrocytes, and the passive behavior and low RM are the intrinsic properties of mature astrocytes. Copyright © 2015, Journal of Neurophysiology.
    Journal of Neurophysiology 03/2015; 113(10):jn.00206.2015. DOI:10.1152/jn.00206.2015 · 2.89 Impact Factor
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    • "Dramatic down-regulation of Kir4.1 channels and their rectification, specifically in M€ uller cells, are associated with vitreo-and chorio-retinal pathology including glaucoma (Francke et al., 1997), retinal detachment (Francke et al., 2005), ischemia (Pannicke et al., 2004, 2005), and diabetes (Pannicke et al., 2006). The Kir channel blocker, spermine, is accumulated in glia rather than in neurons (Biedermann et al., 1998; Laube and Veh, 1997) and localized in the same cellular compartments together with Kir4.1 in M€ uller glial cells (Skatchkov et al., 2000, 2001). However, the amount of SP in the cytoplasm and the interaction between spermine and potassium with Kir4.1 channels remains an enigma. "
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    ABSTRACT: Although Kir4.1 channels are the major inwardly rectifying channels in glial cells and are widely accepted to support K+- and glutamate-uptake in the nervous system, the properties of Kir4.1 channels during vital changes of K+ and polyamines remain poorly understood. Therefore, the present study examined the voltage-dependence of K+ conductance with varying physiological and pathophysiological external [K+] and intrapipette spermine ([SP]) concentrations in Müller glial cells and in tsA201 cells expressing recombinant Kir4.1 channels. Two different types of [SP] block were characterized: "fast" and "slow." Fast block was steeply voltage-dependent, with only a low sensitivity to spermine and strong dependence on extracellular potassium concentration, [K+]o. Slow block had a strong voltage sensitivity that begins closer to resting membrane potential and was essentially [K+]o-independent, but with a higher spermine- and [K+]i-sensitivity. Using a modified Woodhull model and fitting i/V curves from whole cell recordings, we have calculated free [SP](in) in Müller glial cells as 0.81 +/- 0.24 mM. This is much higher than has been estimated previously in neurons. Biphasic block properties underlie a significantly varying extent of rectification with [K+] and [SP]. While confirming similar properties of glial Kir and recombinant Kir4.1, the results also suggest mechanisms underlying K+ buffering in glial cells: When [K+]o is rapidly increased, as would occur during neuronal excitation, "fast block" would be relieved, promoting potassium influx to glial cells. Increase in [K+]in would then lead to relief of "slow block," further promoting K+-influx.
    Glia 05/2008; 56(7):775-90. DOI:10.1002/glia.20652 · 6.03 Impact Factor
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    • "The reduction of the sustained inward-current amplitude , as well as of the inward rectification in the preincubated cells, was probably due to the reduction of Kir 4.1-mediated currents by the ATP deficiency (Takumi et al., 1995; Ishii et al., 1997; Kusaka and Puro, 1997) and to the lack of spermine, the molecule responsible for rectification (Biedermann et al., 1998; Baukrowitz et al., 1999; Skatchkov et al., 2000). There is a dramatic loss of spermine/spermidine immunoreactivity in Mü ller cells after metabolic block (S.N. "
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    ABSTRACT: The retinae and brains of larval and adult amphibians survive long-lasting anoxia; this finding suggests the presence of functional K(ATP) channels. We have previously shown with immunocytochemistry studies that retinal glial (Müller) cells in adult frogs express the K(ATP) channel and receptor proteins, Kir6.1 and SUR1, while retinal neurons display Kir6.2 and SUR2A/B (Skatchkov et al., 2001a: NeuroReport 12:1437-1441; Eaton et al., in press: NeuroReport). Using both immunocytochemistry and electrophysiology, we demonstrate the expression of Kir6.1/SUR1 (K(ATP)) channels in adult frog and tadpole Müller cells. Using conditions favoring the activation of K(ATP) channels (i.e., ATP- and spermine-free cytoplasm-dialyzing solution containing gluconate) in Müller cells isolated from both adult frogs and tadpoles, we demonstrate the following. First, using the patch-clamp technique in whole-cell recordings, tolbutamide, a blocker of K(ATP) channels, blocks nearly 100% of the transient and about 30% of the steady-state inward currents and depolarizes the cell membrane by 5-12 mV. Second, inside-out membrane patches display a single-channel inward current induced by gluconate (40 mM) and blocked by ATP (200 microM) at the cytoplasmic side. The channels apparently show two sublevels (each of approximately 27-32 pS) with a total of 85-pS maximal conductance at -80 mV; the open probability follows a two-exponential mechanism. Thus, functional K(ATP) channels, composed of Kir6.1/SUR1, are present in frog Müller cells and contribute a significant part to the whole-cell K+ inward currents in the absence of ATP. Other inwardly rectifying channels, such as Kir4.1 or Kir2.1, may mediate the remaining currents. K(ATP) channels may help maintain glial cell functions during ATP deficiency.
    Glia 06/2002; 38(3):256-67. DOI:10.1002/glia.10073 · 6.03 Impact Factor
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