Developmental Expression of the Small-Conductance Ca2+-Activated Potassium Channel SK2 in the Rat Retina

doi:10.1006/mcne.2000.0956

Molecular and Cellular Neuroscience

Volume 17, Issue 3, March 2001, Pages 514-520

https://doi.org/10.1006/mcne.2000.0956 Get rights and content

Abstract

Small-conductance Ca²⁺-activated potassium (SK) channels are present in most central neurons, where they mediate the afterhyperpolarizations (AHPs) following action potentials. SK channels integrate changes in intracellular Ca²⁺ concentration with membrane potential and thus play an important role in controlling firing pattern and excitability. Here, we characterize the expression pattern of the apamin-sensitive SK subunits, SK2 and SK3, in the developing and adult rat retina using in situ hybridization and immunohistochemistry. The SK2 subunit showed a distinct and developmentally regulated pattern of expression. It appeared during the first postnatal week and located to retinal ganglion cells and to subpopulations of neurons in the inner nuclear layer. These neurons were identified as horizontal cells and dopaminergic amacrine cells by specific markers. In contrast to SK2, the SK3 subunit was detected neither in the developing nor in the adult retina. These results show cell-specific expression of the SK2 subunit in the retina and suggest that this channel underlies the apamin-sensitive AHP currents described in retinal ganglion cells.

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Citation Excerpt :
Ca2+ influx through Ca2+ channels is also likely responsible for gating Ca2+-activated K+ channels (Feigenspan et al., 1998; Xiao et al., 2004). SK2 channels have been identified in DACs (Klöcker et al., 2001). However, application of the SK channel blocker apamin had minimal effect on DAC action potential waveforms or spiking frequency while the BK channel blocker charybdotoxin slightly accelerated spiking and blocked the action potential after-hyperpolarization (Feigenspan et al., 1998).
In this review, we summarize studies investigating the types and distribution of voltage- and calcium-gated ion channels in the different classes of retinal neurons: rods, cones, horizontal cells, bipolar cells, amacrine cells, interplexiform cells, and ganglion cells. We discuss differences among cell subtypes within these major cell classes, as well as differences among species, and consider how different ion channels shape the responses of different neurons. For example, even though second-order bipolar and horizontal cells do not typically generate fast sodium-dependent action potentials, many of these cells nevertheless possess fast sodium currents that can enhance their kinetic response capabilities. Ca²⁺ channel activity can also shape response kinetics as well as regulating synaptic release. The L-type Ca²⁺ channel subtype, Ca_V1.4, expressed in photoreceptor cells exhibits specific properties matching the particular needs of these cells such as limited inactivation which allows sustained channel activity and maintained synaptic release in darkness. The particular properties of K⁺ and Cl⁻ channels in different retinal neurons shape resting membrane potentials, response kinetics and spiking behavior. A remaining challenge is to characterize the specific distributions of ion channels in the more than 100 individual cell types that have been identified in the retina and to describe how these particular ion channels sculpt neuronal responses to assist in the processing of visual information by the retina.
Calcium-activated potassium channel SK1 is widely expressed in the peripheral nervous system and sensory organs of adult zebrafish
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Citation Excerpt :
As far as we know the presence of SK1 in the retina of the teleost has not been reported earlier. In the rat retina all SK ion channels were absent from the photoreceptors layer, although they are present in other retinal cells [16,24]. Further studies are necessaries to elucidate the role of SK1 in fish retina and why the differences with high vertebrates.
Sensory cells contain ion channels involved in the organ-specific transduction mechanisms that convert different types of stimuli into electric energy. Here we focus on small-conductance calcium-activated potassium channel 1 (SK1) which plays an important role in all excitable cells acting as feedback regulators in after-hyperpolarization. This study was undertaken to analyze the pattern of expression of SK1 in the zebrafish peripheral nervous system and sensory organs using RT-PRC, Westernblot and immunohistochemistry. Expression of SK1 mRNA was observed at all developmental stages analyzed (from 10 to 100 days post fertilization, dpf), and the antibody used identified a protein with a molecular weight of 70 kDa, at 100 dpf (regarded to be adult). Cell expressing SK1 in adult animals were neurons of dorsal root and cranial nerve sensory ganglia, sympathetic neurons, sensory cells in neuromasts of the lateral line system and taste buds, crypt olfactory neurons and photoreceptors. Present results report for the first time the expression and the distribution of SK1 in the peripheral nervous system and sensory organs of adult zebrafish, and may contribute to set zebrafish as an interesting experimental model for calcium-activated potassium channels research. Moreover these findings are of potential interest because the potential role of SK as targets for the treatment of neurological diseases and sensory disorders.
An amino acid outside the pore region influences apamin sensitivity in small conductance Ca2+-activated K+ channels
2007, Journal of Biological Chemistry
Small conductance calcium-activated potassium channels (SK, K_Ca) are a family of voltage-independent K⁺ channels with a distinct physiology and pharmacology. The bee venom toxin apamin inhibits exclusively the three cloned SK channel subtypes (SK1, SK2, and SK3) with different affinity, highest for SK2, lowest for SK1, and intermediate for SK3 channels. The high selectivity of apamin made it a valuable tool to study the molecular makeup and function of native SK channels. Three amino acids located in the outer vestibule of the pore are of particular importance for the different apamin sensitivities of SK channels. Chimeric SK1 channels, enabling the homomeric expression of the rat SK1 (rSK1) subunit and containing the core domain (S1–S6) of rSK1, are apamin-insensitive. By contrast, channels formed by the human orthologue human SK1 (hSK1) are sensitive to apamin. This finding hinted at the involvement of regions beyond the pore as determinants of apamin sensitivity, because hSK1 and rSK1 have an identical amino acid sequence in the pore region. Here we investigated which parts of the channels outside the pore region are important for apamin sensitivity by constructing chimeras between apamin-insensitive and -sensitive SK channel subunits and by introducing point mutations. We demonstrate that a single amino acid situated in the extracellular loop between the transmembrane segments S3 and S4 has a major impact on apamin sensitivity. Our findings enabled us to convert the hSK1 channel into a channel that was as sensitive for apamin as SK2, the SK channel with the highest sensitivity.
Regulation of surface localization of the small conductance Ca 2+-activated potassium channel, Sk2, through direct phosphorylation by cAMP-dependent protein kinase
2006, Journal of Biological Chemistry
Small conductance, Ca²⁺-activated voltage-independent potassium channels (SK channels) are widely expressed in diverse tissues; however, little is known about the molecular regulation of SK channel subunits. Direct alteration of ion channel subunits by kinases is a candidate mechanism for functional modulation of these channels. We find that activation of cyclic AMP-dependent protein kinase (PKA) with forskolin (50 μm) causes a dramatic decrease in surface localization of the SK2 channel subunit expressed in COS7 cells due to direct phosphorylation of the SK2 channel subunit. PKA phosphorylation studies using the intracellular domains of the SK2 channel subunit expressed as glutathione S-transferase fusion protein constructs showed that both the amino-terminal and carboxyl-terminal regions are PKA substrates in vitro. Mutational analysis identified a single PKA phosphorylation site within the amino-terminal of the SK2 subunit at serine 136. Mutagenesis and mass spectrometry studies identified four PKA phosphorylation sites: Ser⁴⁶⁵ (minor site) and three amino acid residues Ser⁵⁶⁸, Ser⁵⁶⁹, and Ser⁵⁷⁰ (major sites) within the carboxyl-terminal region. A mutated SK2 channel subunit, with the three contiguous serines mutated to alanines to block phosphorylation at these sites, shows no decrease in surface expression after PKA stimulation. Thus, our findings suggest that PKA phosphorylation of these three sites is necessary for PKA-mediated reorganization of SK2 surface expression.
Maturation of firing pattern in chick vestibular nucleus neurons
2006, Neuroscience
The principal cells of the chick tangential nucleus are vestibular nucleus neurons participating in the vestibuloocular and vestibulocollic reflexes. In birds and mammals, spontaneous and stimulus-evoked firing of action potentials is essential for vestibular nucleus neurons to generate mature vestibular reflex activity. The emergence of spike-firing pattern and the underlying ion channels were studied in morphologically-identified principal cells using whole-cell patch-clamp recordings from brain slices of late-term embryos (embryonic day 16) and hatchling chickens (hatching day 1 and hatching day 5). Spontaneous spike activity emerged around the perinatal period, since at embryonic day 16 none of the principal cells generated spontaneous action potentials. However, at hatching day 1, 50% of the cells fired spontaneously (range, 3 to 32 spikes/s), which depended on synaptic transmission in most cells. By hatching day 5, 80% of the principal cells could fire action potentials spontaneously (range, 5 to 80 spikes/s), and this activity was independent of synaptic transmission and showed faster kinetics than at hatching day 1. Repetitive firing in response to depolarizing pulses appeared in the principal cells starting around embryonic day 16, when <20% of the neurons fired repetitively. However, almost 90% of the principal cells exhibited repetitive firing on depolarization at hatching day 1, and 100% by hatching day 5. From embryonic day 16 to hatching day 5, the gain for evoked spike firing increased almost 10-fold. At hatching day 5, a persistent sodium channel was essential for the generation of spontaneous spike activity, while a small conductance, calcium-dependent potassium current modulated both the spontaneous and evoked spike firing activity. Altogether, these in vitro studies showed that during the perinatal period, the principal cells switched from displaying no spontaneous spike activity at resting membrane potential and generating one spike on depolarization to the tonic firing of spontaneous and evoked action potentials.
The rat retinal ganglion cell in culture: An accessible CNS neurone
2005, Journal of Pharmacological and Toxicological Methods
Retinal ganglion cells are vital for vision, some have intrinsic light sensing properties and in retinal networks display complex computational abilities. Furthermore they are implicated in a very common form of blindness, glaucoma as well some the symptoms of AIDS. Retinal ganglion cells, unlike many neurones of the central nervous system, have a clearly defined physiological role and can be identified in primary cultures with ease. Here we detail the cell culture and electrophysiological methods required to obtain recordings on the voltage-gated and ligand-gated ion currents and channels expressed by these neurones. Information is given on the range of non-ionotropic receptors that are thought to be present on these cells and what role they may have as model systems in the pharmacological and pharmaceutical research environment.

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Regular ArticleDevelopmental Expression of the Small-Conductance Ca2+-Activated Potassium Channel SK2 in the Rat Retina

Abstract

J. Neurol. Sci.

J. Biol. Chem.

Exp. Eye Res.

Semin. Cell. Dev. Biol.

Brain Res. Dev. Brain Res.

Trends Neurosci.

Neuron

Prog. Brain. Res.

Curr. Opin. Neurobiol.

Neuron

Neuron

Spontaneous activity of solitary dopaminergic cells of the retina

J. Neurosci.

Requirement for cholinergic synaptic transmission in the propagation of spontaneous retinal waves

Science

Age-dependent and cell class-specific modulation of retinal ganglion cell bursting activity by GABA

J. Neurosci.

Spontaneous impulse activity of rat retinal ganglion cells in prenatal life

Science

Localization of two calcium binding proteins, calbindin (28 kD) and parvalbumin (12 kD), in the vertebrate retina

J. Comp. Neurol.

Cellular degeneration and synaptogenesis in the developing retina of the rat

J. Comp. Neurol.

Regular Article
Developmental Expression of the Small-Conductance Ca²⁺-Activated Potassium Channel SK2 in the Rat Retina