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Upregulation of T-type Ca2+ channels in long-term diabetes determines increased excitability of a specific type of capsaicin-insensitive DRG neurons.

Duzhyy DE, Viatchenko-Karpinski VY, Khomula EV, Voitenko NV, Belan PV - Mol Pain (2015)

Bottom Line: This upregulation was not accompanied by significant changes in biophysical properties of T-type channels suggesting that a density of functionally active channels was increased.The upregulation of T-type channels resulted in the increased neuronal excitability of these nociceptive neurons revealed by a lower threshold for action potential initiation, prominent afterdepolarizing potentials and burst firing.Capsaicin-insensitive low-pH-sensitive type of DRG neurons shows diabetes-induced upregulation of Cav3.2 subtype of T-type channels.

View Article: PubMed Central - PubMed

Affiliation: Department of General Physiology of the CNS and State Key Laboratory of Molecular and Cellular Biology, Bogomoletz Institute of Physiology of National Academy of Science of Ukraine, 4 Bogomoletz street, 01024, Kyiv, Ukraine. dduzhyy@biph.kiev.ua.

ABSTRACT

Background: Previous studies have shown that increased excitability of capsaicin-sensitive DRG neurons and thermal hyperalgesia in rats with short-term (2-4 weeks) streptozotocin-induced diabetes is mediated by upregulation of T-type Ca(2+) current. In longer-term diabetes (after the 8th week) thermal hyperalgesia is changed to hypoalgesia that is accompanied by downregulation of T-type current in capsaicin-sensitive small-sized nociceptors. At the same time pain symptoms of diabetic neuropathy other than thermal persist in STZ-diabetic animals and patients during progression of diabetes into later stages suggesting that other types of DRG neurons may be sensitized and contribute to pain. In this study, we examined functional expression of T-type Ca(2+) channels in capsaicin-insensitive DRG neurons and excitability of these neurons in longer-term diabetic rats and in thermally hypoalgesic diabetic rats.

Results: Here we have demonstrated that in STZ-diabetes T-type current was upregulated in capsaicin-insensitive low-pH-sensitive small-sized nociceptive DRG neurons of longer-term diabetic rats and thermally hypoalgesic diabetic rats. This upregulation was not accompanied by significant changes in biophysical properties of T-type channels suggesting that a density of functionally active channels was increased. Sensitivity of T-type current to amiloride (1 mM) and low concentration of Ni(2+) (50 μM) implicates prevalence of Cav3.2 subtype of T-type channels in the capsaicin-insensitive low-pH-sensitive neurons of both naïve and diabetic rats. The upregulation of T-type channels resulted in the increased neuronal excitability of these nociceptive neurons revealed by a lower threshold for action potential initiation, prominent afterdepolarizing potentials and burst firing. Sodium current was not significantly changed in these neurons during long-term diabetes and could not contribute to the diabetes-induced increase of neuronal excitability.

Conclusions: Capsaicin-insensitive low-pH-sensitive type of DRG neurons shows diabetes-induced upregulation of Cav3.2 subtype of T-type channels. This upregulation results in the increased excitability of these neurons and may contribute to nonthermal nociception at a later-stage diabetes.

No MeSH data available.


Related in: MedlinePlus

Direct correlation between the AP parameters characterizing neuronal excitability and TCD in the caps−lpH+ DRG neurons. AP parameters were measured at the AP initiation threshold in Tyrode’s solution. TCD was measured in the same neurons at a voltage step from −100 to −50 mV when the external Tyrode’s solution was changed for TEA-Cl, Ba2+-based solution. a Significant correlation between the ADP/AHP area and TCD. Pearson coefficient Pc = 0.78, p < 1.2*10−5. Number of cells: n = 28 from eight rats b Significant correlation between the AP threshold and TCD. Pearson coefficient, Pc = 0.52, p < 0.0015. Number of cells: n = 36 from eight rats
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Fig5: Direct correlation between the AP parameters characterizing neuronal excitability and TCD in the caps−lpH+ DRG neurons. AP parameters were measured at the AP initiation threshold in Tyrode’s solution. TCD was measured in the same neurons at a voltage step from −100 to −50 mV when the external Tyrode’s solution was changed for TEA-Cl, Ba2+-based solution. a Significant correlation between the ADP/AHP area and TCD. Pearson coefficient Pc = 0.78, p < 1.2*10−5. Number of cells: n = 28 from eight rats b Significant correlation between the AP threshold and TCD. Pearson coefficient, Pc = 0.52, p < 0.0015. Number of cells: n = 36 from eight rats

Mentions: First, we evaluated variation of excitability parameters due to variation in T-type current in the caps−lpH+ neurons. Application of T-type channel blocker, Ni2+ (50 μM), onto a subset of the caps−lpH+ neurons having large ADP (and presumably expressing large T-type current [5]) converted the ADP to AHP and increased an AP threshold by 10 mV (Fig 4e-g). The AP threshold trace under Ni2+ application (grey trace) is shifted to the left compared to control (black trace) because of the higher value of injected current needed to reach the higher level of AP threshold (Fig 4e). Ni2+ application led to a nearly total block of T-type current. Peak transient Ba2+ current was inhibited by 93 ± 2 % at a voltage step to −50 mV (Fig 4d), while total Ba2+ current was inhibited by 84 ± 2 % at this voltage step. Almost complete block of T-type current by 50 μM of Ni2+ suggested that the Cav3.2 T-type channels (IC50 = 13 μM [29]) rather than Cav3.1 or Cav3.3 isoforms (IC50 = 250 μM and 216 μM, correspondently [29]) mainly contributed to T-type current in the caps−lpH+ DRG neurons. For a double check of the effect of T-type current block on excitability parameters we applied the other T-type channel blocker, amiloride, which is selective for Cav3.2 T-type channel isoform [30–32] and, to the best of our knowledge, has no significant inhibitory effect on voltage-gated sodium channels at 1 mM concentration. When applied at this concentration, amiloride inhibited Ba2+ current by 76 ± 7 % at a voltage step to −50 mV (Fig 4d). Weaker inhibition of T-type current with amiloride resulted in a lesser decrease of the ADP area and lower increase of the AP threshold, demonstrating correlation between the T-type current inhibition and the excitability parameters (Fig 4h, i). So, the highest decrease of the ADP area (by 1704 mV*ms; Fig 4 f) was observed when T-type current was maximally inhibited by Ni2+ (by 93 %; Fig 5a) while the ADP area was decreased by 1241 mV*ms; Fig 4h) in case of amiloride application when T-type current was inhibited by 76 % (Fig 4d). Analogous relationship was obtained between the APT and TCD (Fig 4d, g, i). Ni2+-induced inhibition of T-type current led to an APT increase by 10 mV (from −55.4 ± 0.7 to −45.5 ± 2.0 mV) (Fig 4g), while amiloride application increased the APT by 7 mV (from −56.3 ± 1.3 to −49.8 ± 2.0 mV) (Fig 4i). A weaker effect of amiloride compared to Ni2+ correlated with its weaker inhibition of T-type current (76 ± 7 % for amiloride compared to 93 ± 2 % for Ni2+ at a voltage step to −50 mV , Fig 4d).Fig. 5


Upregulation of T-type Ca2+ channels in long-term diabetes determines increased excitability of a specific type of capsaicin-insensitive DRG neurons.

Duzhyy DE, Viatchenko-Karpinski VY, Khomula EV, Voitenko NV, Belan PV - Mol Pain (2015)

Direct correlation between the AP parameters characterizing neuronal excitability and TCD in the caps−lpH+ DRG neurons. AP parameters were measured at the AP initiation threshold in Tyrode’s solution. TCD was measured in the same neurons at a voltage step from −100 to −50 mV when the external Tyrode’s solution was changed for TEA-Cl, Ba2+-based solution. a Significant correlation between the ADP/AHP area and TCD. Pearson coefficient Pc = 0.78, p < 1.2*10−5. Number of cells: n = 28 from eight rats b Significant correlation between the AP threshold and TCD. Pearson coefficient, Pc = 0.52, p < 0.0015. Number of cells: n = 36 from eight rats
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4490764&req=5

Fig5: Direct correlation between the AP parameters characterizing neuronal excitability and TCD in the caps−lpH+ DRG neurons. AP parameters were measured at the AP initiation threshold in Tyrode’s solution. TCD was measured in the same neurons at a voltage step from −100 to −50 mV when the external Tyrode’s solution was changed for TEA-Cl, Ba2+-based solution. a Significant correlation between the ADP/AHP area and TCD. Pearson coefficient Pc = 0.78, p < 1.2*10−5. Number of cells: n = 28 from eight rats b Significant correlation between the AP threshold and TCD. Pearson coefficient, Pc = 0.52, p < 0.0015. Number of cells: n = 36 from eight rats
Mentions: First, we evaluated variation of excitability parameters due to variation in T-type current in the caps−lpH+ neurons. Application of T-type channel blocker, Ni2+ (50 μM), onto a subset of the caps−lpH+ neurons having large ADP (and presumably expressing large T-type current [5]) converted the ADP to AHP and increased an AP threshold by 10 mV (Fig 4e-g). The AP threshold trace under Ni2+ application (grey trace) is shifted to the left compared to control (black trace) because of the higher value of injected current needed to reach the higher level of AP threshold (Fig 4e). Ni2+ application led to a nearly total block of T-type current. Peak transient Ba2+ current was inhibited by 93 ± 2 % at a voltage step to −50 mV (Fig 4d), while total Ba2+ current was inhibited by 84 ± 2 % at this voltage step. Almost complete block of T-type current by 50 μM of Ni2+ suggested that the Cav3.2 T-type channels (IC50 = 13 μM [29]) rather than Cav3.1 or Cav3.3 isoforms (IC50 = 250 μM and 216 μM, correspondently [29]) mainly contributed to T-type current in the caps−lpH+ DRG neurons. For a double check of the effect of T-type current block on excitability parameters we applied the other T-type channel blocker, amiloride, which is selective for Cav3.2 T-type channel isoform [30–32] and, to the best of our knowledge, has no significant inhibitory effect on voltage-gated sodium channels at 1 mM concentration. When applied at this concentration, amiloride inhibited Ba2+ current by 76 ± 7 % at a voltage step to −50 mV (Fig 4d). Weaker inhibition of T-type current with amiloride resulted in a lesser decrease of the ADP area and lower increase of the AP threshold, demonstrating correlation between the T-type current inhibition and the excitability parameters (Fig 4h, i). So, the highest decrease of the ADP area (by 1704 mV*ms; Fig 4 f) was observed when T-type current was maximally inhibited by Ni2+ (by 93 %; Fig 5a) while the ADP area was decreased by 1241 mV*ms; Fig 4h) in case of amiloride application when T-type current was inhibited by 76 % (Fig 4d). Analogous relationship was obtained between the APT and TCD (Fig 4d, g, i). Ni2+-induced inhibition of T-type current led to an APT increase by 10 mV (from −55.4 ± 0.7 to −45.5 ± 2.0 mV) (Fig 4g), while amiloride application increased the APT by 7 mV (from −56.3 ± 1.3 to −49.8 ± 2.0 mV) (Fig 4i). A weaker effect of amiloride compared to Ni2+ correlated with its weaker inhibition of T-type current (76 ± 7 % for amiloride compared to 93 ± 2 % for Ni2+ at a voltage step to −50 mV , Fig 4d).Fig. 5

Bottom Line: This upregulation was not accompanied by significant changes in biophysical properties of T-type channels suggesting that a density of functionally active channels was increased.The upregulation of T-type channels resulted in the increased neuronal excitability of these nociceptive neurons revealed by a lower threshold for action potential initiation, prominent afterdepolarizing potentials and burst firing.Capsaicin-insensitive low-pH-sensitive type of DRG neurons shows diabetes-induced upregulation of Cav3.2 subtype of T-type channels.

View Article: PubMed Central - PubMed

Affiliation: Department of General Physiology of the CNS and State Key Laboratory of Molecular and Cellular Biology, Bogomoletz Institute of Physiology of National Academy of Science of Ukraine, 4 Bogomoletz street, 01024, Kyiv, Ukraine. dduzhyy@biph.kiev.ua.

ABSTRACT

Background: Previous studies have shown that increased excitability of capsaicin-sensitive DRG neurons and thermal hyperalgesia in rats with short-term (2-4 weeks) streptozotocin-induced diabetes is mediated by upregulation of T-type Ca(2+) current. In longer-term diabetes (after the 8th week) thermal hyperalgesia is changed to hypoalgesia that is accompanied by downregulation of T-type current in capsaicin-sensitive small-sized nociceptors. At the same time pain symptoms of diabetic neuropathy other than thermal persist in STZ-diabetic animals and patients during progression of diabetes into later stages suggesting that other types of DRG neurons may be sensitized and contribute to pain. In this study, we examined functional expression of T-type Ca(2+) channels in capsaicin-insensitive DRG neurons and excitability of these neurons in longer-term diabetic rats and in thermally hypoalgesic diabetic rats.

Results: Here we have demonstrated that in STZ-diabetes T-type current was upregulated in capsaicin-insensitive low-pH-sensitive small-sized nociceptive DRG neurons of longer-term diabetic rats and thermally hypoalgesic diabetic rats. This upregulation was not accompanied by significant changes in biophysical properties of T-type channels suggesting that a density of functionally active channels was increased. Sensitivity of T-type current to amiloride (1 mM) and low concentration of Ni(2+) (50 μM) implicates prevalence of Cav3.2 subtype of T-type channels in the capsaicin-insensitive low-pH-sensitive neurons of both naïve and diabetic rats. The upregulation of T-type channels resulted in the increased neuronal excitability of these nociceptive neurons revealed by a lower threshold for action potential initiation, prominent afterdepolarizing potentials and burst firing. Sodium current was not significantly changed in these neurons during long-term diabetes and could not contribute to the diabetes-induced increase of neuronal excitability.

Conclusions: Capsaicin-insensitive low-pH-sensitive type of DRG neurons shows diabetes-induced upregulation of Cav3.2 subtype of T-type channels. This upregulation results in the increased excitability of these neurons and may contribute to nonthermal nociception at a later-stage diabetes.

No MeSH data available.


Related in: MedlinePlus