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Inorganic polyphosphate regulates neuronal excitability through modulation of voltage-gated channels.

Stotz SC, Scott LO, Drummond-Main C, Avchalumov Y, Girotto F, Davidsen J, Gómez-Gárcia MR, Rho JM, Pavlov EV, Colicos MA - Mol Brain (2014)

Bottom Line: Mechanistically, this is accomplished by shifting the voltage sensitivity of NaV channel activation toward the neuronal resting membrane potential, the block KV channels, and the activation of CaV channels.Next, using calcium imaging we found that polyP stimulates an increase in neuronal network activity and induces calcium influx in glial cells.We conclude that polyP release leads to increased excitability of the neuronal membrane through the modulation of voltage gated ion channels.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physiology & Pharmacology and the Hotchkiss Brain Institute, University of Calgary, 3330 Hospital Drive NW, Calgary, AB T2N 4N1, Canada. mcolicos@ucalgary.ca.

ABSTRACT

Background: Inorganic polyphosphate (polyP) is a highly charged polyanion capable of interacting with a number of molecular targets. This signaling molecule is released into the extracellular matrix by central astrocytes and by peripheral platelets during inflammation. While the release of polyP is associated with both induction of blood coagulation and astrocyte extracellular signaling, the role of secreted polyP in regulation of neuronal activity remains undefined. Here we test the hypothesis that polyP is an important participant in neuronal signaling. Specifically, we investigate the ability of neurons to release polyP and to induce neuronal firing, and clarify the underlying molecular mechanisms of this process by studying the action of polyP on voltage gated channels.

Results: Using patch clamp techniques, and primary hippocampal and dorsal root ganglion cell cultures, we demonstrate that polyP directly influences neuronal activity, inducing action potential generation in both PNS and CNS neurons. Mechanistically, this is accomplished by shifting the voltage sensitivity of NaV channel activation toward the neuronal resting membrane potential, the block KV channels, and the activation of CaV channels. Next, using calcium imaging we found that polyP stimulates an increase in neuronal network activity and induces calcium influx in glial cells. Using in situ DAPI localization and live imaging, we demonstrate that polyP is naturally present in synaptic regions and is released from the neurons upon depolarization. Finally, using a biochemical assay we demonstrate that polyP is present in synaptosomes and can be released upon their membrane depolarization by the addition of potassium chloride.

Conclusions: We conclude that polyP release leads to increased excitability of the neuronal membrane through the modulation of voltage gated ion channels. Together, our data establishes that polyP could function as excitatory neuromodulator in both the PNS and CNS.

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Related in: MedlinePlus

Neuronal response to polyP. Both CNS and PNS neurons responded to the application of polyP with a vehement burst of activity. A) Representative response of a hippocampal neuron to direct application of polyP (n = 16 of 24), in which rapid, persistent firing is observed. B) Representative response of a DRG neuron to polyP. Small nociceptive DRG neurons (<35 microns) responded with rapid firing (n = 24). C) Some neurons responded to polyP with a multiphasic response: rapid firing followed by protracted depolarization, during which time firing did not occur, that was restored upon recovery of the membrane potential (8 of 24).
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Figure 1: Neuronal response to polyP. Both CNS and PNS neurons responded to the application of polyP with a vehement burst of activity. A) Representative response of a hippocampal neuron to direct application of polyP (n = 16 of 24), in which rapid, persistent firing is observed. B) Representative response of a DRG neuron to polyP. Small nociceptive DRG neurons (<35 microns) responded with rapid firing (n = 24). C) Some neurons responded to polyP with a multiphasic response: rapid firing followed by protracted depolarization, during which time firing did not occur, that was restored upon recovery of the membrane potential (8 of 24).

Mentions: Current-clamp recordings of cultured hippocampal and DRG neurons determined that polyP can functionally stimulate excitable cells. Puff application of medium chain length polyP molecules (60 orthophosphate groups in length on average, see Methods for concentration details) elicited a dramatic increase in AP firing rates from both the CNS (Figure 1A) and PNS (Figure 1B) neurons. Additionally, some hippocampal neurons (n = 8 of 24) had a multi-phasic response to polyP: AP firing frequency increased initially, but ceased as the membrane resting potential depolarized, then AP firing reappeared as the neurons repolarized (Figure 1C). The polyP concentration was chosen based on the reported average polyP concentrations observed in CNS tissue [9,10].


Inorganic polyphosphate regulates neuronal excitability through modulation of voltage-gated channels.

Stotz SC, Scott LO, Drummond-Main C, Avchalumov Y, Girotto F, Davidsen J, Gómez-Gárcia MR, Rho JM, Pavlov EV, Colicos MA - Mol Brain (2014)

Neuronal response to polyP. Both CNS and PNS neurons responded to the application of polyP with a vehement burst of activity. A) Representative response of a hippocampal neuron to direct application of polyP (n = 16 of 24), in which rapid, persistent firing is observed. B) Representative response of a DRG neuron to polyP. Small nociceptive DRG neurons (<35 microns) responded with rapid firing (n = 24). C) Some neurons responded to polyP with a multiphasic response: rapid firing followed by protracted depolarization, during which time firing did not occur, that was restored upon recovery of the membrane potential (8 of 24).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4061113&req=5

Figure 1: Neuronal response to polyP. Both CNS and PNS neurons responded to the application of polyP with a vehement burst of activity. A) Representative response of a hippocampal neuron to direct application of polyP (n = 16 of 24), in which rapid, persistent firing is observed. B) Representative response of a DRG neuron to polyP. Small nociceptive DRG neurons (<35 microns) responded with rapid firing (n = 24). C) Some neurons responded to polyP with a multiphasic response: rapid firing followed by protracted depolarization, during which time firing did not occur, that was restored upon recovery of the membrane potential (8 of 24).
Mentions: Current-clamp recordings of cultured hippocampal and DRG neurons determined that polyP can functionally stimulate excitable cells. Puff application of medium chain length polyP molecules (60 orthophosphate groups in length on average, see Methods for concentration details) elicited a dramatic increase in AP firing rates from both the CNS (Figure 1A) and PNS (Figure 1B) neurons. Additionally, some hippocampal neurons (n = 8 of 24) had a multi-phasic response to polyP: AP firing frequency increased initially, but ceased as the membrane resting potential depolarized, then AP firing reappeared as the neurons repolarized (Figure 1C). The polyP concentration was chosen based on the reported average polyP concentrations observed in CNS tissue [9,10].

Bottom Line: Mechanistically, this is accomplished by shifting the voltage sensitivity of NaV channel activation toward the neuronal resting membrane potential, the block KV channels, and the activation of CaV channels.Next, using calcium imaging we found that polyP stimulates an increase in neuronal network activity and induces calcium influx in glial cells.We conclude that polyP release leads to increased excitability of the neuronal membrane through the modulation of voltage gated ion channels.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physiology & Pharmacology and the Hotchkiss Brain Institute, University of Calgary, 3330 Hospital Drive NW, Calgary, AB T2N 4N1, Canada. mcolicos@ucalgary.ca.

ABSTRACT

Background: Inorganic polyphosphate (polyP) is a highly charged polyanion capable of interacting with a number of molecular targets. This signaling molecule is released into the extracellular matrix by central astrocytes and by peripheral platelets during inflammation. While the release of polyP is associated with both induction of blood coagulation and astrocyte extracellular signaling, the role of secreted polyP in regulation of neuronal activity remains undefined. Here we test the hypothesis that polyP is an important participant in neuronal signaling. Specifically, we investigate the ability of neurons to release polyP and to induce neuronal firing, and clarify the underlying molecular mechanisms of this process by studying the action of polyP on voltage gated channels.

Results: Using patch clamp techniques, and primary hippocampal and dorsal root ganglion cell cultures, we demonstrate that polyP directly influences neuronal activity, inducing action potential generation in both PNS and CNS neurons. Mechanistically, this is accomplished by shifting the voltage sensitivity of NaV channel activation toward the neuronal resting membrane potential, the block KV channels, and the activation of CaV channels. Next, using calcium imaging we found that polyP stimulates an increase in neuronal network activity and induces calcium influx in glial cells. Using in situ DAPI localization and live imaging, we demonstrate that polyP is naturally present in synaptic regions and is released from the neurons upon depolarization. Finally, using a biochemical assay we demonstrate that polyP is present in synaptosomes and can be released upon their membrane depolarization by the addition of potassium chloride.

Conclusions: We conclude that polyP release leads to increased excitability of the neuronal membrane through the modulation of voltage gated ion channels. Together, our data establishes that polyP could function as excitatory neuromodulator in both the PNS and CNS.

Show MeSH
Related in: MedlinePlus