ABSTRACT
In the present study, ATP-activated currents (I(ATP)s) recorded from rat trigeminal ganglion (TG) neurons using whole-cell patch clamp technique are classified into three types (F, I and S) based on the characteristics of their activation and desensitization. The time of rising phase (R(10-90)) of types F, I and S of I(ATP) is measured to be 33.6+/-4.5, 62.2+/-9.9 and 302.1+/-62.0 ms respectively, and positively correlated to cell size. The time of decaying phases (D(10-90)) of types F and S is 399.4+/-58.2 and >1500 ms, respectively. The dose-response curves for the three types of I(ATP) show that their EC(50) values are close (3.44 x 10(-5), 4.89 x 10(-5) and 4.14 x 10(-5) M for types F, I and S respectively, P>0.05). Their reversal potentials are basically the same, varying from +4 to +10 mV. In addition, using whole-cell patch clamp technique in combination with single cell immunohistochemical staining for P2X receptor subunits, our results suggest that the type distinction of ATP-activated current was associated with cell size and P2X receptor subunits: small-sized cells with type F of I(ATP) express only P2X1 and/or P2X3 subunits, while cells with types S and I of I(ATP) express P2X2 or P2X4 in addition to P2X1 and P2X3.
Subject(s)
Adenosine Triphosphate/metabolism , Neurons, Afferent/metabolism , Receptors, Purinergic P2/metabolism , Trigeminal Ganglion/metabolism , Adenosine Triphosphate/agonists , Adenosine Triphosphate/analogs & derivatives , Animals , Animals, Newborn , Cell Membrane/drug effects , Cell Membrane/metabolism , Cells, Cultured , Dose-Response Relationship, Drug , Immunohistochemistry , Membrane Potentials/drug effects , Membrane Potentials/physiology , Neurons, Afferent/drug effects , Patch-Clamp Techniques , Protein Subunits/drug effects , Protein Subunits/metabolism , Purinergic P2 Receptor Agonists , Purinergic P2 Receptor Antagonists , Rats , Rats, Sprague-Dawley , Receptors, Purinergic P2/chemistry , Receptors, Purinergic P2X , Receptors, Purinergic P2X2 , Receptors, Purinergic P2X3 , Receptors, Purinergic P2X4 , Trigeminal Ganglion/drug effectsABSTRACT
The characteristics of purinoceptors in the membrane of rat trigeminal ganglion (TG) neurons were studied by using whole- cell patch clamp technique. The results showed that most of neurons examined (78.9%, 142/180) were responsive to ATP in a concentration-dependent manner; the others (21.1%, 38/180) were ATP insensitive. Of the ATP-sensitive cells, the majority (95.1%, 135/142) responded to ATP with an inward current, a few (2.1%, 3/142) with an outward current, and the rest (2.8%, 4/142) with biphasic current. Small sized cells (<30 mum) responded to ATP with a rapid desensitizing inward current and were highly sensitive to vanilloid; the medium sized cells (30~40 mum) responded to ATP with slow desensitizing inward current and were not sensitive to vanilloid; while the majority of large sized cells (>40 mum) did not respond to ATP and vanilloid. The waveform of ATP-activated inward currents was related to the cell diameter. The I-V curves for both small and medium sized cells manifested obvious inward rectification. Furthermore, we studied the kinetic features of ATP-activated currents and the effects of P2 purinoceptor agonists and antagonists on I(ATP). The findings suggest that ATP receptor-ion channels are expressed differently among different types of rat TG neurons.
Subject(s)
Adenosine Triphosphate/metabolism , Neurons/metabolism , Receptors, Purinergic P2X/physiology , Trigeminal Ganglion/metabolism , Animals , Animals, Newborn , Neurons/physiology , Patch-Clamp Techniques , Rats , Rats, Sprague-Dawley , Trigeminal Ganglion/cytology , Trigeminal Ganglion/physiologyABSTRACT
In response to SP applied externally, neurons freshly isolated from bullfrog dorsal root ganglion (DRG) showed three kinds of current (I(SP)), i.e. slow, fast and moderately activating I(SP)s. All the three kinds of I(SP) were inward currents, and were completely blocked by either peptide antagonist of SP receptor spantide or non-peptide antagonist of SP receptor WIN51708. The slow activating I(SP) showed slow kinetic features. Replacement of NaCl in external solution by NMDG had no effect on this kind of I(SP), while Ba(2+) abolished it almost completely, thus the ionic mechanism underlying slow activating I(SP) was deduced to be the closure of K(+) channels. The fast activating I(SP) in bullfrog DRG neurons, just as in rat DRG neurons, was proved to be caused by the opening of Na(+) preferring non-selective cation channel, for it was abolished almost completely by replacement of NaCl in external solution with equimolar NMDG. The moderately activating I(SP) was similar to the fast activating I(SP) in current configuration, however, its kinetic characteristics lay between those of fast and slow activating I(SP)s. Either NMDG or Ba(2+) suppressed this kind of I(SP) partially. Therefore the moderately activating I(SP) might be mediated by non-selective cation channel. We used repatch technique to explore the intracellular mechanism underlying the three kinds of I(SP) and found that the three kinds of I(SP) were caused by the activity of either G-protein coupled channel (slow activating I(SP)) or directly opened channel (fast activating I(SP)) or both (moderately activating I(SP) ).
