Calcium signaling in intact dorsal root ganglia: new observations and the effect of injury.
BACKGROUND
Ca is the dominant second messenger in primary sensory neurons. In addition, disrupted Ca signaling is a prominent feature in pain models involving peripheral nerve injury. Standard cytoplasmic Ca recording techniques use high K or field stimulation and dissociated neurons. To compare findings in intact dorsal root ganglia, we used a method of simultaneous electrophysiologic and microfluorimetric recording.


METHODS
Dissociated neurons were loaded by bath-applied Fura-2-AM and subjected to field stimulation. Alternatively, we adapted a technique in which neuronal somata of intact ganglia were loaded with Fura-2 through an intracellular microelectrode that provided simultaneous membrane potential recording during activation by action potentials (APs) conducted from attached dorsal roots.


RESULTS
Field stimulation at levels necessary to activate neurons generated bath pH changes through electrolysis and failed to predictably drive neurons with AP trains. In the intact ganglion technique, single APs produced measurable Ca transients that were fourfold larger in presumed nociceptive C-type neurons than in nonnociceptive Abeta-type neurons. Unitary Ca transients summated during AP trains, forming transients with amplitudes that were highly dependent on stimulation frequency. Each neuron was tuned to a preferred frequency at which transient amplitude was maximal. Transients predominantly exhibited monoexponential recovery and had sustained plateaus during recovery only with trains of more than 100 APs. Nerve injury decreased Ca transients in C-type neurons, but increased transients in Abeta-type neurons.


CONCLUSIONS
Refined observation of Ca signaling is possible through natural activation by conducted APs in undissociated sensory neurons and reveals features distinct to neuronal types and injury state.
Gemes
Geza
G
Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA.
Rigaud
Marcel
M
Koopmeiners
Andrew S
AS
Poroli
Mark J
MJ
Zoga
Vasiliki
V
Hogan
Quinn H
QH
D000200:Action Potentials; D000818:Animals; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D020013:Calcium Signaling; D003592:Cytophotometry / Q000379:methods; D016257:Fura-2 / Q000008:administration & dosage / Q000031:analogs & derivatives; D005727:Ganglia, Spinal / Q000293:injuries* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008297:Male; D008564:Membrane Potentials; D009412:Nerve Fibers; D009475:Neurons, Afferent; D051381:Rats; D017207:Rats, Sprague-Dawley; D011984:Sensory Receptor Cells / Q000378:metabolism*
D002121:Calcium Channel Blockers; D015220:Calcium Channels; C049925:fura-2-am; D016257:Fura-2