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Neurons were Stimulated Using a Piezo-Electric Controlled Heat-Polished Glass Pipette

Neurons were Stimulated Using a Piezo-Electric Controlled Heat-Polished Glass Pipette. Perforated Patch Configuration Neurons 1 day in culture Tests: Series 2 m increments Repeated stimulus. Characterisation of Mechanically Activated Currents in Dorsal Root Ganglia Neurons.

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Neurons were Stimulated Using a Piezo-Electric Controlled Heat-Polished Glass Pipette

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  1. Neurons were Stimulated Using a Piezo-Electric Controlled Heat-Polished Glass Pipette • Perforated Patch • Configuration • Neurons 1 day • in culture • Tests: • Series 2m increments • Repeated stimulus

  2. Characterisation of Mechanically Activated Currents in Dorsal Root Ganglia Neurons • LJ Drew, TM Jessell, JN Wood & • P Cesare

  3. Caps - Rapidly adapting Capsaicin Sensitive and Insensitive Neurons are Differentially Responsive to Mechanical Stimulation Stimuli 12 mm 0 Inward Currents 3000 0 2000 Current Amplitude (pA) -3000 0 1000 pA 0 -600 0 2 4 6 8 10 12 0 Displacement (mm) -500 Caps - Slowly adapting 0 300 Caps + Rapidly adapting Time (msec)

  4. Blockade of MA Currents by Ruthenium Red and Gadolinium in Different Subpopulations 100 100 % Inhibition 50 50 0 0 0.1 1 10 100 0.1 1 10 100 Log10 [Gd3+] (mM) Log10 [Ruthenium Red] (mM) Caps + Rapidly adapting Caps - Rapidly adapting Caps - Slowly adapting

  5. MA Currents in Subsets of DRG Neurons are Differentially Modulated by Ca2+ Caps - 0mM Ca2+ 0 +100 pA -500 +50 5mM Ca2+ -1000 % Control (2mM Ca2+) 0 Caps + 0 -50 -400 pA Caps - 5mM Ca2+ -100 Caps + Control -800 0mM Ca2+

  6. MA Currents Are Acutely Inhibited by Cytochalasin B (10M) in Capsaicin Insensitive Neurons P = 0.0001 100 80 60 % Control 40 20 0 Caps + Caps -

  7. Are MA Currents ASIC Mediated? • ASIC2 and ASIC3 KO Mice show moderate changes in mechanosensitivity. • Magnitude of MA currents does not correlate with rapidly adapting low pH evoked currents. • MA currents are not modulated by low pH. • MA currents are not sensitive to Amiloride. • MA currents are blocked by 300M Zn2+.

  8. Conclusions • Important aspects of the mechanosensitive phenotype of DRG neurons are maintained in vitro. • Pharmacology: MA Currents are mediated by closely related ion channels in low- and high-threshold neurons. • Ca2+ modulation reveals diversity in MA current modulation. • Sensitivity of ion channels may be determined by cytoskeletal tethering.

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