This study explored whether KMUP-1 improved chronic constriction injury (CCI)-induced BKCa current inhibition in dorsal root ganglion (DRG) neurons. Rats were randomly assigned to four groups: sham, ...sham + KMUP-1, CCI, and CCI + KMUP-1 (5 mg/kg/day, i.p.). DRG neuronal cells (L4–L6) were isolated on day 7 after CCI surgery. Perforated patch-clamp and inside-out recordings were used to monitor BKCa currents and channel activities, respectively, in the DRG neurons. Additionally, DRG neurons were immunostained with anti-NeuN, anti-NF200 and anti-BKCa. Real-time PCR was used to measure BKCa mRNA levels. In perforated patch-clamp recordings, CCI-mediated nerve injury inhibited BKCa currents in DRG neurons compared with the sham group, whereas KMUP-1 prevented this effect. CCI also decreased BKCa channel activity, which was recovered by KMUP-1 administration. Immunofluorescent staining further demonstrated that CCI reduced BKCa-channel proteins, and KMUP-1 reversed this. KMUP-1 also changed CCI-reduced BKCa mRNA levels. KMUP-1 prevented CCI-induced neuropathic pain and BKCa current inhibition in a peripheral nerve injury model, suggesting that KMUP-1 could be a potential agent for controlling neuropathic pain.
Abstract Every 23 s, a person sustains a traumatic brain injury in the United States leaving many patients with substantial cognitive impairment and epilepsy. Injury-induced alterations in the ...hippocampus underpin many of these disturbances of neurological function. Abnormalities in the dentate gyrus are likely to play a major role in the observed pathophysiology because this subregion functions as a filter impeding excessive or aberrant activity from propagating further into the circuit and following experimental brain injury, the dentate gyrus becomes more excitable. Although alteration in excitation or inhibition could mediate this effect in the dentate gyrus, we show a key role played by an impairment of GABAA ergic inhibition. The efficacy of GABAA -mediated inhibition depends on a low Cl− i that is maintained by neuronal K–Cl co-transporter 2 (KCC2). Using fluid percussion injury (FPI) in the mouse, we demonstrate significant reductions in KCC2 protein and mRNA expression in the dentate gyrus that causes a depolarizing shift in GABAA reversal potential, due to impaired chloride clearance, resulting in reduced inhibitory efficiency. This study elucidates a novel mechanism underlying diminished dentate gyrus inhibitory efficacy and provides an innovative target for the development of potential therapeutics to restore the severe pathological consequences of traumatic brain injury.
Background:
Inflammation-induced sensitization of primary afferents is associated with a decrease in K+ current. However, the type of K+ current and basis for the decrease varies as a function of ...target of innervation. Because glabrous skin of the rat hindpaw is used often to assess changes in nociception in models of persistent pain, the purpose of the present study was to determine the type and extent to which K+ currents contribute to the inflammation-induced sensitization of cutaneous afferents. Acutely dissociated retrogradely labeled cutaneous dorsal root ganglion neurons from naïve and inflamed (3 days post complete Freund's adjuvant injection) rats were studied with whole cell and perforated patch techniques.
Results:
Inflammation-induced sensitization of small diameter cutaneous neurons was associated with an increase in action potential duration and rate of decay of the afterhyperpolarization. However, no changes in voltage-gated K+ currents were detected. In contrast, Ca2+ modulated iberiotoxin sensitive and paxilline sensitive K+ (BKCa) currents were significantly smaller in small diameter IB4+ neurons. This decrease in current was not associated with a detectable change in total protein levels of the BKCa channel α or β subunits. Single cell PCR analysis revealed a significant change in the pattern of expression of α subunit splice variants and β subunits that were consistent, at least in part, with inflammation-induced changes in the biophysical properties of BKCa currents in cutaneous neurons.
Conclusions:
Results of this study provide additional support for the conclusion that it may be possible, if not necessary to selectively treat pain arising from specific body regions. Because a decrease in BKCa current appears to contribute to the inflammation-induced sensitization of cutaneous afferents, BKCa channel openers may be effective for the treatment of inflammatory pain.
Modulation of the membrane excitability of rat parasympathetic intracardiac ganglion neurons by muscarinic receptors was studied using an amphotericin B-perforated patch-clamp recording ...configuration. Activation of muscarinic receptors by oxotremorine-M (OxoM) depolarized the membrane, accompanied by repetitive action potentials. OxoM evoked inward currents under voltage-clamp conditions at a holding potential of −60 mV. Removal of extracellular Ca2+ markedly increased the OxoM-induced current (IOxoM). The inward IOxoM in the absence of extracellular Ca2+ was fully inhibited by removal of extracellular Na+, indicating the involvement of non-selective cation channels. The IOxoM was inhibited by organic cation channel antagonists including SKF-96365 and ML-204. The IOxoM was antagonized by muscarinic receptor antagonists with the following potency: 4-DAMP > pirenzepine = darifenacin > methoctramine. Muscarinic toxin 7 (MT-7), a highly selective inhibitor for M1 receptor, produced partial inhibition of the IOxoM. In the presence of MT-7, concentration–inhibition curve of the M3-preferring antagonist darifenacin was shifted to the left. These results suggest the contribution of M1 and M3 receptors to the OxoM response. The IOxoM was inhibited by U-73122, a phospholipase C inhibitor. The membrane-permeable IP3 receptor blocker xestospongin C also inhibited the IOxoM. Furthermore, pretreatment with thapsigargin and BAPTA-AM inhibited the IOxoM, while KN-62, a blocker of Ca2+/calmodulin-dependent protein kinase II, had no effect. These results suggest that the activation mechanism involves a PLC pathway, release of Ca2+ from intracellular Ca2+ stores and calmodulin.
The cation channels activated by muscarinic receptors may play an important role in neuronal membrane depolarization in rat intracardiac ganglion neurons.
•Role of the muscarinic receptor in intracardiac ganglion neurons was investigated.•Muscarinic receptor agonist oxotremorine-M excited the ganglion neurons.•Oxotremorine-M induced cation currents via M1 and M3 receptors.•The oxotremorine-M response was mediated by intracellular Ca2+ release.•The results suggest that muscarinic receptors contribute to ganglionic transmission.
Many mammalian central nervous system neuron responses mediated by GABA(A) receptors undergo a developmental transition from excitation to inhibition, but little is known about the time of this ...switch in specific cell types in the developing anteroventral cochlear nucleus (AVCN). In the present study, bushy and stellate cells, two major cell types in the AVCN, were identified according to their morphology and electrophysiology. The equilibrium potential of GABA-evoked currents (E(GABA)) was examined using the gramicidin-perforated patch-clamp technique. We found that the action of GABA in bushy and stellate cells switched from predominantly depolarizing to predominantly hyperpolarizing with respect to their resting membrane potential (V(rest)) at different postnatal ages. Such a switch in the GABA response of bushy cells occurred before the first postnatal week, whereas that in stellate cells happened at the end of the second postnatal week. Furthermore, we discovered that bushy cells had a more depolarized V(rest) than did stellate cells before the second postnatal week; however, the E(GABA) of bushy and stellate cells was not significantly different. Thus, the discrepancy in the timing of the developmental shift from depolarizing to hyperpolarizing GABA responses between bushy and stellate cells may be due to the difference in their V(rest), but not due to E(GABA) itself. These results suggest that GABAergic inhibition functions earlier in bushy than in stellate cells. In contrast, the longer excitatory action of GABA on stellate cells possibly renders them more vulnerable than bushy cells to excitotoxic substances during early development.
Abstract Lidocaine hydrochloride (LC-HCl) is widely used as a local anesthetic, while various adverse effects of LC-HCl, such as seizures have also been reported. Lidocaine is reported to inhibit ...various channels and receptors including GABAA receptors. Although the GABAA receptor-mediated response depends on Cl− equilibrium potential ( ECl ), little is known about the effect of LC-HCl on ECl . In the present study, we investigated the effect of LC-HCl on GABA-induced currents in cultured rat hippocampal neurons with gramicidin-perforated patch–clamp recording which is known to keep the intracellular Cl− concentration intact. LC-HCl inhibited outward GABA-induced currents with depolarizing shift of the GABA reversal potential ( EGABA ). The LC-HCl-induced positive EGABA shift was not observed with conventional whole-cell patch–clamp method which cannot retain intact intracellular Cl− concentration. The LC-HCl action on EGABA was inhibited by either furosemide, a blocker of both Na+ -K+ -Cl− cotransporter (NKCC) and K+ -Cl− cotransporter (KCC), or an increase in extracellular K+ concentrations. Neither bumetanide, a specific inhibitor of NKCC, nor Na+ -free external solution had any effect on the LC-HCl-induced EGABA shift. QX-314, a membrane impermeable lidocaine derivative, failed to shift EGABA to positive potential. Furthermore, LC-HCl caused a depolarizing shift of EGABA in cultured GT1-7 cells expressing KCC2 but failed to change EGABA in GT1-7 cells without expression of KCC2. These results suggest that the LC-HCl-induced positive EGABA shift is due to a blockade of KCC2. Together with the direct LC-HCl action to GABAA receptors, the positive EGABA shift induced by LC-HCl reduces the GABAergic inhibition in the central nervous system.
Coronoids, patches and generalised altans Bašić, Nino; Fowler, Patrick W.; Pisanski, Tomaž
Journal of mathematical chemistry,
04/2016, Letnik:
54, Številka:
4
Journal Article
Recenzirano
Odprti dostop
In this paper we revisit coronoids, in particular multiple coronoids. We consider a mathematical formalisation of the theory of coronoid hydrocarbons that is solely based on incidence between ...hexagons of the infinite hexagonal grid in the plane. In parallel, we consider perforated patches, which generalise coronoids: in addition to hexagons, other polygons may also be present. Just as coronoids may be considered as benzenoids with holes, perforated patches are patches with holes. Both cases, coronoids and perforated patches, admit a generalisation of the altan operation that can be performed at several holes simultaneously. A formula for the number of Kekulé structures of a generalised altan can be derived easily if the number of Kekulé structures is known for the original graph. Pauling Bond Orders for generalised altans are also easy to derive from those of the original graph.
Electrical stimulation (ES) of skeletal muscle partially mimics the benefits of physical activity. However, the stimulation protocols applied clinically to date, often cause unpleasant symptoms and ...muscle fatigue. Here, we compared the efficiency of a “noisy” stimulus waveform derived from human electromyographic (EMG) muscle patterns, with stereotyped 45 and 1 Hz electrical stimulations applied to mouse myotubes in vitro. Human gastrocnemius medialis electromyograms recorded from volunteers during real locomotor activity were used as a template for a noisy stimulation, called EMG
stim
. The stimulus-induced electrical activity, intracellular Ca
2+
dynamics and mechanical twitches in the myotubes were assessed using whole-cell perforated patch-clamp, Ca
2+
imaging and optical visualization techniques. EMG
stim
was more efficient in inducing myotube cell firing, Ca
2+
i
changes and contractions compared with more conventional electrical stimulation. Its stimulation strength was also much lower than the minimum required to induce contractions via stereotyped stimulation protocols. We conclude that muscle cells in vitro can be more efficiently depolarized using the “noisy” stochastic stimulation pattern, EMG
stim
, a finding that suggests a way to favor a higher level of electrical activity in a larger number of cells.
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