Precise control of ion channel and gap junction expression is required for patterning of the regenerating axolotl limb

Axolotls and other salamanders have the capacity to regenerate lost tissue after an amputation or injury. Growth and morphogenesis are coordinated within cell groups in many contexts by the interplay of transcriptional networks and biophysical properties such as ion flows and voltage gradients. It is not, however, known whether regulators of a cell’s ionic state are involved in limb patterning at later stages of regeneration. Here we manipulated expression and activities of ion channels and gap junctions in vivo, in axolotl limb blastema cells. Limb amputations followed by retroviral infections were performed to drive expression of a human gap junction protein Connexin 26 (Cx26), potassium (Kir2.1-Y242F and Kv1.5) and sodium (NeoNav1.5) ion channel proteins along with EGFP control. Skeletal preparation revealed that overexpressing Cx26 caused syndactyly, while overexpression of ion channel proteins resulted in digit loss and structural abnormalities compared to EGFP expressing control limbs. Additionally, we showed that exposing limbs to the gap junction inhibitor lindane during the regeneration process caused digit loss. Our data reveal that manipulating native ion channel and gap junction function in blastema cells results in patterning defects involving the number and structure of the regenerated digits. Gap junctions and ion channels have been shown to mediate ion flows that control the endogenous voltage gradients which are tightly associated with the regulation of gene expression, cell cycle progression, migration, and other cellular behaviors. Therefore, we postulate that mis-expression of these channels may have disturbed this regulation causing uncoordinated cell behavior which results in morphological defects.

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WHAT CAUSES ION CHANNEL PROTEINS TO FLUCTUATE OPEN AND CLOSED?

International Journal of Neural Systems ◽

10.1142/s0129065796000282 ◽

1996 ◽

Vol 07 (04) ◽

pp. 321-331 ◽

Cited By ~ 10

Author(s):

LARRY S. LIEBOVITCH ◽

ANGELO T. TODOROV

Keyword(s):

Ion Channels ◽

Patch Clamp ◽

Ion Channel ◽

Patch Clamp Technique ◽

Sodium Potassium ◽

Random Event ◽

Channel Protein ◽

Channel Proteins ◽

Ion Channel Proteins ◽

Open States

Ion channels in the cell membrane spontaneously switch from states that are closed to the flow of ions such as sodium, potassium, and chloride to states that are open to the flow of these ions. The durations of times that an individual ion channel protein spends in the closed and open states can be measured by the patch clamp technique. We explore two basic issues about the molecular properties of ion channels: 1) If the switching between the closed and open state is an inherently random event, what does the patch clamp data tell us about the structure or motions in the ion channel protein? 2) Is this switching random?

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Ion channels and disease

Oxford Textbook of Medicine ◽

10.1093/med/9780198746690.003.0032 ◽

2020 ◽

pp. 246-255

Author(s):

Frances Ashcroft ◽

Paolo Tammaro

Keyword(s):

Ion Channels ◽

Ion Channel ◽

Single Channel ◽

Cell Types ◽

Channel Structure ◽

Channel Proteins ◽

Channel Function ◽

Ion Channel Proteins ◽

Single Channel Currents ◽

Channel Currents

Ion channels are membrane proteins that act as gated pathways for the movement of ions across cell membranes. They are found in both surface and intracellular membranes and play essential roles in the physiology of all cell types. An ever-increasing number of human diseases are now known to be caused by defects in ion channel function. To understand how ion channel defects give rise to disease, it is helpful to understand how the ion channel proteins work. This chapter therefore considers what is known of ion channel structure, explains the properties of the single ion channel, and shows how single-channel currents give rise to action potentials and synaptic potentials.

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Regulation of ion channel structure and function by reactive oxygen-nitrogen species

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00281.2003 ◽

2003 ◽

Vol 285 (6) ◽

pp. L1184-L1189 ◽

Cited By ~ 63

Author(s):

Sadis Matalon ◽

Karin M. Hardiman ◽

Lucky Jain ◽

Douglas C. Eaton ◽

Michael Kotlikoff ◽

...

Keyword(s):

Gene Expression ◽

Ion Channels ◽

Ion Channel ◽

Posttranslational Modifications ◽

Regulation Of Gene Expression ◽

Reactive Oxygen ◽

Cellular Level ◽

Channel Structure ◽

Nitrogen Species ◽

Channel Proteins

Ion channels subserve diverse cellular functions. Reactive oxygen and nitrogen species modulate ion channel function by a number of mechanisms including 1) transcriptional regulation of gene expression, 2) posttranslational modifications of channel proteins, i.e. nitrosylation, nitration, and oxidation of key amino acid residues, 3) by altering the gain in other signaling pathways that may in turn lead to changes in channel activity or channel gene expression, and 4) by modulating trafficking or turnover of channel proteins, as typified by oxygen radical activation of NF-kB, with subsequent changes in proteasomal degradation of channel degradation. Regardless of the mechanism, as was discussed in a symposium at the 2003 Experimental Biology Meeting in San Diego, CA, changes in the cellular level of reactive oxygen and nitrogen species can have profound effects on the activity of ion channels and cellular function.

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Ion channel proteins in mouse and human vestibular tissue

Otolaryngology ◽

10.1016/j.otohns.2005.01.022 ◽

2005 ◽

Vol 132 (6) ◽

pp. 916-923 ◽

Cited By ~ 4

Author(s):

Karin Hotchkiss ◽

Margaret Harvey ◽

Mary Pacheco ◽

Bernd Sokolowski

Keyword(s):

Ion Channels ◽

Ion Channel ◽

Inner Ear ◽

Potassium Ion ◽

Β Subunit ◽

Potassium Ion Channel ◽

Channel Proteins ◽

Ion Channel Proteins ◽

Vestibular Endorgans ◽

Α Subunits

BACKGROUND AND OBJECTIVE: Electrical activity in hair cells and neurons of the inner ear is necessary for the transduction and modulation of stimuli that impinge on the cochlea and vestibular endorgans of the inner ear. The underlying basis of this activity is pore-forming proteins in the membrane of excitable cells that allow the influx and efflux of various ions, including Na+, Ca2+, and K+, among others. These channels are critical to both electrical activity as well as the development of excitable cells because they may initiate long-term signals that are important in the maintenance and survival of these cells. We investigated the expression of several Shaker potassium ion channel proteins and an accessory β subunit in the vestibular endorgans of mouse and human. METHODS: Vestibular tissue consisting of cristae ampullares was harvested from adult and neonatal mice as well as from human subjects undergoing vestibular surgery. Western blot analysis and immunoprecipitation were used to identify the presence or absence, in mouse, of α subunits Kv1.2, Kv1.4, and Kv1.5 and of β subunit Kvβ1.1 in mouse. Coimmunoprecipitation was used to identify interactions between α and β subunits. Immunohistochemistry was used to localize Kv1.2 in mouse and human tissues. RESULTS: The presence of Kvα1.2 and Kvβ1.1 was confirmed in adult mouse crista ampullaris by Western blotting. Coimmunoprecipitation experiments showed that Kv1.2 and Kvβ1.1 interact in these tissues. Immunostaining localized Kv1.2 to regions within and extraneous to the sensory epithelium of mouse and human cristae ampullares. In comparison, Kv1.4 and Kv1.5 were not found in the crista ampullaris. CONCLUSIONS: We describe the presence, location, and interaction of various potassium ion channel α subunits and a β subunit. These data are initial descriptions of potassium ion channels in the mammalian vestibular system and begin to provide an understanding of the protein subunits that form ion channels of the mammalian inner ear. In addition, our data show that there are interactions that occur that may regulate the biophysical properties of these channels, thereby contributing to the diversity of channel function. This knowledge is critical to understanding the genes that encode these channels and finding cures for pathologies of hearing and balance. SIGNIFICANCE: We detail initial characteristics of potassium ion channel proteins including α subunits Kv1.2, Kv1.4, and Kv1.5 and β subunit Kvβ1.1 in mammalian vestibular tissue. This knowledge is critical to understanding the processing of vestibular stimuli and the regulation of endolymphatic function. Mutations of ion channels can cause neurological pathologies including auditory and vestibular disorders in humans.

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