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The chemokine CXCL1/growth related oncogene increases sodium currents and neuronal excitability in small diameter sensory neurons.

Wang JG, Strong JA, Xie W, Yang RH, Coyle DE, Wick DM, Dorsey ED, Zhang JM - Mol Pain (2008)

Bottom Line: These effects required long exposures, and were completely blocked by co-incubation with protein synthesis inhibitor cycloheximide.Many studies on the role of chemokines in pain conditions have focused on their rapid and indirect effects on neurons, via release of inflammatory mediators from immune and glial cells.Our study suggests that GRO/KC may also have important pro-nociceptive effects via its direct actions on sensory neurons, and may induce long-term changes that involve protein synthesis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Anesthesiology, University of Cincinnati College of Medicine, Cincinnati, OH 45267-0531, USA. jungang.wang@uchsc.edu

ABSTRACT

Background: Altered Na+ channel expression, enhanced excitability, and spontaneous activity occur in nerve-injury and inflammatory models of pathological pain, through poorly understood mechanisms. The cytokine GRO/KC (growth related oncogene; CXCL1) shows strong, rapid upregulation in dorsal root ganglion in both nerve injury and inflammatory models. Neurons and glia express its receptor (CXCR2). CXCL1 has well-known effects on immune cells, but little is known about its direct effects on neurons.

Results: We report that GRO/KC incubation (1.5 nM, overnight) caused marked upregulation of Na+ currents in acutely isolated small diameter rat (adult) sensory neurons in vitro. In both IB4-positive and IB4-negative sensory neurons, TTX-resistant and TTX-sensitive currents increased 2- to 4 fold, without altered voltage dependence or kinetic changes. These effects required long exposures, and were completely blocked by co-incubation with protein synthesis inhibitor cycloheximide. Amplification of cDNA from the neuronal cultures showed that 3 Na channel isoforms were predominant both before and after GRO/KC treatment (Nav 1.1, 1.7, and 1.8). TTX-sensitive isoforms 1.1 and 1.7 significantly increased 2 - 3 fold after GRO/KC incubation, while 1.8 showed a trend towards increased expression. Current clamp experiments showed that GRO/KC caused a marked increase in excitability, including resting potential depolarization, decreased rheobase, and lower action potential threshold. Neurons acquired a striking ability to fire repetitively; IB4-positive cells also showed marked broadening of action potentials. Immunohistochemical labelling confirmed that the CXCR2 receptor was present in most neurons both in dissociated cells and in DRG sections, as previously shown for neurons in the CNS.

Conclusion: Many studies on the role of chemokines in pain conditions have focused on their rapid and indirect effects on neurons, via release of inflammatory mediators from immune and glial cells. Our study suggests that GRO/KC may also have important pro-nociceptive effects via its direct actions on sensory neurons, and may induce long-term changes that involve protein synthesis.

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Overnight incubation with GRO/KC does not affect decay time constants or activation of Na+ currents. A. Activation data for the TTX-R current was fit by a standard Boltzmann equation. No differences were found in the fit parameters between the 4 experimental groups (p = 0.22). The plotted line represents the best fit to all the data, and has a V1/2 value of -7.3 mV and a slope factor of 5.7. B. Time constants for the decay of TTX-R current were obtained by fitting single exponentials to the falling phase of currents evoked from a holding potential of -50 mV. No significant difference between the two groups was observed (two-way RM ANOVA). C. Time constants for the TTX-S current at were obtained by fitting the decaying phase of the current with the sum of two exponentials. The slower of these corresponded to the time constant observed in the TTX-R current, and the faster time constant was used as the value of the TTX-S decay time constant. No significant difference between the two groups was observed (Mann-Whitney test, p = 0.18). Data from IB4-positive and IB4-negative cells were combined as no difference between these groups in decay time constants was observed.
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Figure 3: Overnight incubation with GRO/KC does not affect decay time constants or activation of Na+ currents. A. Activation data for the TTX-R current was fit by a standard Boltzmann equation. No differences were found in the fit parameters between the 4 experimental groups (p = 0.22). The plotted line represents the best fit to all the data, and has a V1/2 value of -7.3 mV and a slope factor of 5.7. B. Time constants for the decay of TTX-R current were obtained by fitting single exponentials to the falling phase of currents evoked from a holding potential of -50 mV. No significant difference between the two groups was observed (two-way RM ANOVA). C. Time constants for the TTX-S current at were obtained by fitting the decaying phase of the current with the sum of two exponentials. The slower of these corresponded to the time constant observed in the TTX-R current, and the faster time constant was used as the value of the TTX-S decay time constant. No significant difference between the two groups was observed (Mann-Whitney test, p = 0.18). Data from IB4-positive and IB4-negative cells were combined as no difference between these groups in decay time constants was observed.

Mentions: The voltage dependence of activation for the TTX-resistant current was not significantly affected by GRO/KC incubation (Figure 3A) and was not different between IB4-positive and IB4-negative cells. The activation curve for the combined data was well fit by a Boltzmann equation with V1/2 = -7.3 mV and a slope factor of 5.7. The time course of decay for the TTX-R current during depolarizing pulses was also unaffected by GRO/KC (Figure 3B). Similarly, the time constant for decay of the TTX-S current at -10 mV was not affected by GRO/KC incubation (Figure 3C). The TTX-sensitive current became too fast to accurately voltage clamp at more positive potentials, so its activation curve was not determined in these experiments. However, the TTX-sensitive current observed during steps from -80 to -10 mV or below, where it is readily kinetically separated from the TTX-resistant current (see Methods), showed a qualitatively similar threshold for activation after GRO/KC incubation (see Figure 2C and 2D).


The chemokine CXCL1/growth related oncogene increases sodium currents and neuronal excitability in small diameter sensory neurons.

Wang JG, Strong JA, Xie W, Yang RH, Coyle DE, Wick DM, Dorsey ED, Zhang JM - Mol Pain (2008)

Overnight incubation with GRO/KC does not affect decay time constants or activation of Na+ currents. A. Activation data for the TTX-R current was fit by a standard Boltzmann equation. No differences were found in the fit parameters between the 4 experimental groups (p = 0.22). The plotted line represents the best fit to all the data, and has a V1/2 value of -7.3 mV and a slope factor of 5.7. B. Time constants for the decay of TTX-R current were obtained by fitting single exponentials to the falling phase of currents evoked from a holding potential of -50 mV. No significant difference between the two groups was observed (two-way RM ANOVA). C. Time constants for the TTX-S current at were obtained by fitting the decaying phase of the current with the sum of two exponentials. The slower of these corresponded to the time constant observed in the TTX-R current, and the faster time constant was used as the value of the TTX-S decay time constant. No significant difference between the two groups was observed (Mann-Whitney test, p = 0.18). Data from IB4-positive and IB4-negative cells were combined as no difference between these groups in decay time constants was observed.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2562993&req=5

Figure 3: Overnight incubation with GRO/KC does not affect decay time constants or activation of Na+ currents. A. Activation data for the TTX-R current was fit by a standard Boltzmann equation. No differences were found in the fit parameters between the 4 experimental groups (p = 0.22). The plotted line represents the best fit to all the data, and has a V1/2 value of -7.3 mV and a slope factor of 5.7. B. Time constants for the decay of TTX-R current were obtained by fitting single exponentials to the falling phase of currents evoked from a holding potential of -50 mV. No significant difference between the two groups was observed (two-way RM ANOVA). C. Time constants for the TTX-S current at were obtained by fitting the decaying phase of the current with the sum of two exponentials. The slower of these corresponded to the time constant observed in the TTX-R current, and the faster time constant was used as the value of the TTX-S decay time constant. No significant difference between the two groups was observed (Mann-Whitney test, p = 0.18). Data from IB4-positive and IB4-negative cells were combined as no difference between these groups in decay time constants was observed.
Mentions: The voltage dependence of activation for the TTX-resistant current was not significantly affected by GRO/KC incubation (Figure 3A) and was not different between IB4-positive and IB4-negative cells. The activation curve for the combined data was well fit by a Boltzmann equation with V1/2 = -7.3 mV and a slope factor of 5.7. The time course of decay for the TTX-R current during depolarizing pulses was also unaffected by GRO/KC (Figure 3B). Similarly, the time constant for decay of the TTX-S current at -10 mV was not affected by GRO/KC incubation (Figure 3C). The TTX-sensitive current became too fast to accurately voltage clamp at more positive potentials, so its activation curve was not determined in these experiments. However, the TTX-sensitive current observed during steps from -80 to -10 mV or below, where it is readily kinetically separated from the TTX-resistant current (see Methods), showed a qualitatively similar threshold for activation after GRO/KC incubation (see Figure 2C and 2D).

Bottom Line: These effects required long exposures, and were completely blocked by co-incubation with protein synthesis inhibitor cycloheximide.Many studies on the role of chemokines in pain conditions have focused on their rapid and indirect effects on neurons, via release of inflammatory mediators from immune and glial cells.Our study suggests that GRO/KC may also have important pro-nociceptive effects via its direct actions on sensory neurons, and may induce long-term changes that involve protein synthesis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Anesthesiology, University of Cincinnati College of Medicine, Cincinnati, OH 45267-0531, USA. jungang.wang@uchsc.edu

ABSTRACT

Background: Altered Na+ channel expression, enhanced excitability, and spontaneous activity occur in nerve-injury and inflammatory models of pathological pain, through poorly understood mechanisms. The cytokine GRO/KC (growth related oncogene; CXCL1) shows strong, rapid upregulation in dorsal root ganglion in both nerve injury and inflammatory models. Neurons and glia express its receptor (CXCR2). CXCL1 has well-known effects on immune cells, but little is known about its direct effects on neurons.

Results: We report that GRO/KC incubation (1.5 nM, overnight) caused marked upregulation of Na+ currents in acutely isolated small diameter rat (adult) sensory neurons in vitro. In both IB4-positive and IB4-negative sensory neurons, TTX-resistant and TTX-sensitive currents increased 2- to 4 fold, without altered voltage dependence or kinetic changes. These effects required long exposures, and were completely blocked by co-incubation with protein synthesis inhibitor cycloheximide. Amplification of cDNA from the neuronal cultures showed that 3 Na channel isoforms were predominant both before and after GRO/KC treatment (Nav 1.1, 1.7, and 1.8). TTX-sensitive isoforms 1.1 and 1.7 significantly increased 2 - 3 fold after GRO/KC incubation, while 1.8 showed a trend towards increased expression. Current clamp experiments showed that GRO/KC caused a marked increase in excitability, including resting potential depolarization, decreased rheobase, and lower action potential threshold. Neurons acquired a striking ability to fire repetitively; IB4-positive cells also showed marked broadening of action potentials. Immunohistochemical labelling confirmed that the CXCR2 receptor was present in most neurons both in dissociated cells and in DRG sections, as previously shown for neurons in the CNS.

Conclusion: Many studies on the role of chemokines in pain conditions have focused on their rapid and indirect effects on neurons, via release of inflammatory mediators from immune and glial cells. Our study suggests that GRO/KC may also have important pro-nociceptive effects via its direct actions on sensory neurons, and may induce long-term changes that involve protein synthesis.

Show MeSH
Related in: MedlinePlus