A delayed rectifier current is modulated by the circadian pacemaker in Bulla

Basal retinal neurons of the marine mollusc Bulla gouldiana continue to express a circadian modulation of their membrane conductance for at least two cycles in cell culture. Voltage-dependent currents of these pacemaker cells were recorded using the whole-cell perforated patch-clamp technique to characterize outward currents and investigate their putative circadian modulation. Three components of the outward potassium current were identified. A transient outward current (IA) was activated after depolarization from holding potentials greater than -30 mV, inactivated with a time constant of 50 ms, and partially blocked by 4-aminopyridine (1-5 mM). A Ca(2+)-dependent potassium current (IK(Ca)) was activated by depolarization to potentials more positive than -10 mV and was blocked by removing Ca2+ from the bath or by applying the Ca2+ channel blockers Cd2+ (0.1-0.2 mM) and Ni2+ (1-5 mM). A sustained Ca(2+)-independent current component including the delayed rectifier current (IK) was recorded at potentials positive to -20 mV in the absence of extracellular Na+ and Ca2+ and was partially blocked by tetraethylammonium chloride (TEA, 30mM). Whole-cell currents recorded before and after the projected dawn and normalized to the cell capacitance revealed a circadian modulation of the delayed rectifier current (IK). However, the IA and IK(Ca) currents were not affected by the circadian pacemaker.     

大鼠发育过程中下丘脑神经元钾离子通道温度效应变化的研究

为了探讨出生后钾离子通道在下丘脑神经元热敏感分化过程中的作用,采用膜片钳技术研究出生一个月内SD大鼠急性分离神经元的温度效应,结果表明IK电流密度在出生后一个月内变化不大(P>0.05),而IA电流密度则呈现为升高趋势(P<0.05).同时升高温度,不同出生日期的钾通道NPo都有不同程度的升高,但相较P1d的神经元来说,温度对P18d的电压依赖性影响更大一些.同时温度对IK和IA的影响是不一样的,IA的Q10>2,所有这些显示IA通道在神经元温度敏感性的发育分化过程中起着重要的作用.     

Ionic conductances in two types of sensory neurons in the leech, Macrobdella decora

1. Ion conductances were investigated in two kinds of leech sensory neurons (PM and N1 cells) which differ in their membrane excitability and action potential. 2. In the PM cell body excitable membrane behavior is dominated by only two currents, a sodium current (INa) and a delayed rectifier (IK). 3. In contrast, in the N1 cell INa and IK is supplemented with the presence of a transient potassium current, IA. 4. A comparison between INa and IK in the two cell types did not reveal any significant difference in activation and inactivation kinetics of either current between neurons. 5. Thus, the properties and presence of the A-current in the N1 cell and not in the PM cell may account for the functional difference in excitability of the two kinds of neurons.     

海马神经细胞胆固醇含量降低对其电压依赖钾电流的上调作用

胆固醇普遍存在于细胞膜中,其含量在细胞增殖、生长及各种疾病条件下会发生改变,这暗示胆固醇对细胞功能的调节起着重要的作用。运用全细胞膜片钳技术研究了胆固醇含量变化对海马神经细胞电压依赖钾电流的影响。实验观察到神经细胞经胆固醇去除剂β-甲基环化糊精(MβCD)处理后,胆固醇含量的减少促进了延迟整流钾电流IK的增加,且延缓了瞬间失活钾电流IA的失活。更进一步,延迟整流钾电流IK和瞬间失活钾电流IA分别经TEA和4-AP阻断后,MβCD对两种电流成分的影响显著降低。这一结果进一步表明胆固醇去除剂对电压依赖钾电流的上调是通过作用于IK和IA电流而共同实现的。基于电压依赖钾通道在神经细胞功能中的重要作用,实验结果暗示神经细胞胆固醇含量变化可对神经细胞的兴奋性起调节作用。     

Inactivation of A currents and A channels on rat nodose neurons in culture

Cultured sensory neurons from nodose ganglia were investigated with whole-cell patch-clamp techniques and single-channel recordings to characterize the A current. Membrane depolarization from -40 mV holding potential activated the delayed rectifier current (IK) at potentials positive to -30 mV; this current had a sigmoidal time course and showed little or no inactivation. In most neurons, the A current was completely inactivated at the -40 mV holding potential and required hyperpolarization to remove the inactivation; the A current was isolated by subtracting the IK evoked by depolarizations from -40 mV from the total outward current evoked by depolarizations from -90 mV. The decay of the A current on several neurons had complex kinetics and was fit by the sum of three exponentials whose time constants were 10-40 ms, 100-350 ms, and 1-3 s. At the single-channel level we found that one class of channel underlies the A current. The conductance of A channels varied with the square root of the external K concentration: it was 22 pS when exposed to 5.4 mM K externally, the increased to 40 pS when exposed to 140 mM K externally. A channels activated rapidly upon depolarization and the latency to first opening decreased with depolarization. The open time distributions followed a single exponential and the mean open time increased with depolarization. A channels inactivate in three different modes: some A channels inactivated with little reopening and gave rise to ensemble averages that decayed in 10-40 ms; other A channels opened and closed three to four times before inactivating and gave rise to ensemble averages that decayed in 100-350 ms; still other A channels opened and closed several hundred times and required seconds to inactivate. Channels gating in all three modes contributed to the macroscopic A current from the whole cell, but their relative contribution differed among neurons. In addition, A channels could go directly from the closed, or resting, state to the inactivated state without opening, and the probability for channels inactivating in this way was greater at less depolarized voltages. In addition, a few A channels appeared to go reversibly from a mode where inactivation occurred rapidly to a slow mode of inactivation.     

Stem cells from the adult human brain develop into functional neurons in culture

Recent research communications indicate that the adult human brain contains undifferentiated, multipotent precursors or neural stem cells. It is not known, however, whether these cells can develop into fully functional neurons. We cultured cells from the adult human ventricular wall as neurospheres and passed them at the individual cell level to secondary neurospheres. Following dissociation and plating, the cells developed the antigen profile of the three main cell types in the brain (GFAP, astrocytes; O2, oligodendrocytes; and beta-III-tubulin/NeuN, neurons). More importantly, the cells developed the electrophysiological profiles of neurons and glia. Over a period of 3 weeks, neuron-like cells went through the same phases as neurons do during development in vivo, including up-regulation of inward Na+ -currents, drop in input resistance, shortening of the action potential, and hyperpolarization of the cell membrane. The cells developed overshooting action potentials with a mature configuration. Recordings in voltage-clamp mode displayed both the fast inactivating TTX-sensitive sodium current (INa) underlying the rising phase of the action potential and the two potassium currents terminating the action potential in mature neurons (IA and IK, sensitive to 4-AP and TEA, respectively). We have thus demonstrated that the human ventricular wall contains multipotent cells that can differentiate into functionally mature neurons.     

A slow voltage-activated potassium current in rat vagal neurons.

Potassium currents play a key role in controlling the excitability of neurons. In this paper we describe the properties of a novel voltage-activated potassium current in neurons of the rat dorsal motor nucleus of the vagus (DMV). Intracellular recordings were made from DMV neurons in transverse slices of the medulla. Under voltage clamp, depolarization of these neurons from hyperpolarized membrane potentials (more negative than -80 mV) activated two transient outward currents. One had fast kinetics and had properties similar to A-currents. The other current had an activation threshold of around -95 mV (from a holding potential -110 mV) and inactivated with a time constant of about 3s. It had a reversal potential close to the potassium equilibrium potential. This current was not calcium dependent and was not blocked by 4-aminopyridine (5 mM), catechol (5 mM) or tetraethylammonium (20 mM). It was completely inactivated at the resting membrane potential. This current therefore represents a new type of voltage-activated potassium current. It is suggested that this current might act as a brake to repetitive firing when the neuron is depolarized from membrane potentials negative to the resting potential.     

Alteration of the transmembrane K+ gradient during development of delayed rectifier in isolated rat pulmonary arterial cells

The properties of the tail current associated with the delayed rectifier K+ current (IK) in isolated rat pulmonary artery smooth muscle cells were examined using the whole cell patch clamp technique. The tail currents observed upon repolarization to -60 mV after brief (e.g., 20 ms) or small (i.e. to potentials negative of 0 mV) depolarizations were outwardly directed, as expected given the calculated K+ reversal potential of -83 mV. The tail currents seen upon repolarization after longer (e.g., 500 ms) and larger (e.g., to +60 mV) depolarizations tended to be inwardly directed. Depolarizations of intermediate strength and/or duration were followed by biphasic tail currents, which were inwardly directed immediately upon repolarization, but changed direction and became outwardly directed before deactivation was complete. When cells were depolarized to +60 mV for 500 ms both IK and the subsequent inward tail current at -60 mV were similarly blocked by phencyclidine. Both IK and the inward tail current were also blocked by 4-aminopyridine. Application of progressively more depolarized 30 s preconditioning potentials inactivated IK, and reduced the inward tail current amplitude with a similar potential dependency. These results indicated that the inward tail current was mediated by IK. The reversal potential of the tail current became progressively more positive with longer depolarizations to +60 mV, shifting from -76.1 +/- 2.2 mV (n = 10) after a 20-ms step to -57.7 +/- 3.5 mV (n = 9) after a 500-ms step. Similar effects occurred when extracellular K+ and Na+ were replaced by choline. When extracellular K+ was raised to 50 mM, the tail current was always inwardly directed at -60 mV, but showed little change in amplitude as the duration of depolarization was increased. These observations are best explained if the dependencies of tail current direction and kinetics upon the duration of the preceding depolarization result from an accumulation of K+ at the external face of the membrane, possibly in membrane invaginations. A mathematical model which simulates the reversal potential shift and the biphasic kinetics of the tail current on this basis is presented.     

Properties of transient K+ currents and underlying single K+ channels in rat olfactory receptor neurons

The transient potassium current, IK(t), of enzymatically dissociated rat olfactory receptor neurons was studied using patch-clamp techniques. Upon depolarization from negative holding potentials, IK(t) activated rapidly and then inactivated with a time course described by the sum of two exponential components with time constants of 22.4 and 143 ms. Single-channel analysis revealed a further small component with a time constant of several seconds. Steady-state inactivation was complete at -20 mV and completely removed at -80 mV (midpoint -45 mV). Activation was significant at -40 mV and appeared to reach a maximum conductance at +40 mV (midpoint -13 mV). Deactivation was described by the sum of two voltage-dependent exponential components. Recovery from inactivation was extraordinarily slow (50 s at -100 mV) and the underlying processes appeared complex. IK(t) was reduced by 4-aminopyridine and tetraethylammonium applied externally. Increasing the external K+ concentration ([K+]o) from 5 to 25 mM partially removed IK(t) inactivation, usually without affecting activation kinetics. The elevated [K+]o also hyperpolarized the steady-state inactivation curve by 9 mV and significantly depolarized the voltage dependence of activation. Single transient K+ channels, with conductances of 17 and 26 pS, were observed in excised patches and often appeared to be localized into large clusters. These channels were similar to IK(t) in their kinetic, pharmacological, and voltage-dependent properties and their inactivation was also subject to modulation by [K+]o. The properties of IK(t) imply a role in action potential repolarization and suggest it may also be important in modulating spike parameters during neuronal burst firing. A simple method is also presented to correct for errors in the measurement of whole-cell resistance (Ro) that can result when patch-clamping very small cells. The analysis revealed a mean corrected Ro of 26 G omega for these cells.     

Interaction of barium ions with potassium channels in squid giant axons.

Blocking of potassium channels by internally and externally applied barium ions has been studied in squid giant axons. Internal Ba (3-5 mM) causes rapid decay or "inactivation" of potassium current (IK). The kinetics and degree of block are strongly voltage-dependent. Large positive voltages speed blocking and make it more profound. Raising the external potassium concentration (Ko) from 0 to 250 mM has the opposite effect: block is made slower and less severe. In contrast, for positive voltages block by the tetraethylammonium derivative 3-phenylpropyltriethylammonium ion is almost independent of Ko and voltage. Recovery from block by internal Ba has a rapid phase lasting a few milliseconds and a slow phase lasting approximately 5 min. Internal Ba causes a "hook" in the IK tails recorded on repolarizing the fiber in high potassium external medium. External Ba, on the other hand, blocks without much altering IK time-course. KD (the dissociation constant) for block by external Ba is a few millimolar, and depends on the internal potassium concentration, the holding potential, and other factors. A reaction scheme for Ba and K channels is presented, postulating that internal and external Ba reach the same point in the channel. Once there, Ba blocks and also stabilizes the closed conformation of the channel. The extreme stability of the Ba channel complex implies the existence of negative charge within the channel.     

Neurons differentiating from murine neural crest in culture exhibit sensory or sympathetic-like calcium currents.

The trunk neural crest gives rise to peripheral sensory and sympathetic neurons. In culture, neural crest cells can be induced to differentiate into either neuronal phenotype. Few studies have examined the differentiation of physiological properties in cultures of neural crest cells. Using whole-cell recordings, our study examined the effects of growth factors on high-voltage-activated calcium current profiles exhibited by neurons differentiating in culture. We compared these profiles with those exhibited by sensory and sympathetic neurons. Neural crest cells in culture gave rise to neurons with calcium current profiles identical to either sensory or sympathetic neurons, depending on the growth conditions. On average, the calcium current profile for sensory neurons was 23% (L), 51% (N), and 12% (P), while sympathetic neurons had a similar L-type current (20%), higher N-type (76%), and lower P-type (4%). Neural crest cells cultured with human leukemia inhibitory factor plus somite cells produced neurons with a sympathetic-like calcium current profile (L: 17%, N: 75%, and P: 4%). However, murine leukemia inhibitory factor (L: 25%, N: 52%, and P: 13%) and ciliary neurotrophic factor (L: 18%, N: 49%, and P: 9%) plus somite cells produced neurons with sensory-like calcium current profiles. These growth conditions did not modify the calcium current profiles of neurons cultured from embryonic and neonatal ganglia. Similarly, murine leukemia inhibitory factor produced a greater percentage of neurons (57%) with sensitivity to capsaicin (sensory phenotype) than human leukemia inhibitory factor (3%). Physiological traits can be a useful tool for the determination of neuronal phenotype in culture where other traits may be less stable.     

Regulation of intrinsic excitability in hippocampal neurons by activity-dependent modulation of the KV2.1 potassium channel

K_V2.1 is the prominent somatodendritic sustained or delayed rectifier voltage-gated potassium (Kv) channel in mammalian central neurons, and is a target for activity-dependent modulation via calcineurin-dependent dephosphorylation. Using hanatoxin-mediated block of K_V2.1 we show that, in cultured rat hippocampal neurons, glutamate stimulation leads to significant hyperpolarizing shifts in the voltage-dependent activation and inactivation gating properties of the K_V2.1-component of delayed rectifier K⁺ (IK) currents. In computer models of hippocampal neurons, these glutamate-stimulated shifts in the gating of the K_V2.1-component of IK lead to a dramatic suppression of action potential firing frequency. Current-clamp experiments in cultured rat hippocampal neurons showed glutamate-stimulation induced a similar suppression of neuronal firing frequency. Membrane depolarization also resulted in similar hyperpolarizing shifts in the voltage-dependent gating properties of neuronal IK currents, and suppression of neuronal firing. The glutamate-induced effects on neuronal firing were eliminated by hanatoxin, but not by dendrotoxin-K, a blocker of K_V1.1-containing channels. These studies together demonstrate a specific contribution of modulation of K_V2.1 channels in the activity-dependent regulation of intrinsic neuronal excitability.     

Voltage-gated transient currents in bovine adrenal fasciculata cells. II. A-type K+ current

In whole cell patch clamp recordings on enzymatically dissociated adrenal zona fasciculata (AZF) cells, a rapidly inactivating A-type K+ current was observed in each of more than 150 cells. Activation of IA was steeply voltage dependent and could be described by a Boltzmann function raised to an integer power of 4, with a midpoint of -28.3 mV. Using the "limiting logarithmic potential sensitivity," the single channel gating charge was estimated to be 7.2 e. Voltage-dependent inactivation could also be described by a Boltzmann function with a midpoint of -58.7 mV and a slope factor of 5.92 mV. Gating kinetics of IA included both voltage-dependent and -independent transitions in pathways between closed, open, and inactivated states. IA activated with voltage-dependent sigmoidal kinetics that could be fit with an n4h formalism. The activation time constant, tau a, reached a voltage- independent minimum at potentials positive to 0 mV. IA currents inactivated with two time constants that were voltage independent at potentials ranging from -30 to +45 mV. At +20 mV, tau i(fast) and tau i(slow) were 13.16 +/- 0.64 and 62.26 +/- 5.35 ms (n = 34), respectively. In some cells, IA inactivation kinetics slowed dramatically after many minutes of whole cell recording. Once activated by depolarization, IA channels returned to the closed state along pathways with two voltage-dependent time constants which were 0.208 s, tau rec-f and 10.02 s, tau rec-s at -80 mV. Approximately 90% of IA current recovered with slow kinetics at potentials between -60 and -100 mV. IA was blocked by 4-aminopyridine (IC50 = 629 microM) through a mechanism that was strongly promoted by channel activation. Divalent and trivalent cations including Ni2+ and La3+ also blocked IA with IC50's of 467 and 26.4 microM, respectively. With respect to biophysical properties and pharmacology, IA in AZF cells resembles to some extent transient K+ currents in neurons and muscle, where they function to regulate action potential frequency and duration. The function of this prominent current in steroid hormone secretion by endocrine cells that may not generate action potentials is not yet clear.     

二氧化硫衍生物对大鼠海马CA3区神经元瞬间外向钾电流的影响

为研究二氧化硫(SO2)衍生物——NaHSO3和Na2SO3(二者分子比为1：3)对大鼠海马CA3区神经元瞬间外向钾电流(IA)的影响,利用全细胞膜片钳技术,根据动力学和药理学特性分离鉴定大鼠海马CA3区神经元IA,观察SO2衍生物对IA的效应。发现SO2代谢衍生物可浓度依赖性地增大IA,使IA增大50％的剂量为25μmol/L。此外还与电压呈依赖关系,但不具有频率依赖性。10μmol/L的SO2代谢衍生物不影响IA电流的激活过程,但升高了A-通道稳态失活电压,延长了A-电流失活时间。说明SO2代谢衍生物可增大大鼠海马CA3区神经元IA电流,延长A-电流的失活时间,从而影响海马神经元的膜生理感应,这可能是SO2影响神经细胞功能的机理之一。     

白介素1β时间依赖性地抑制大鼠皮层神经元钾电流

白细胞介素1β（Interleukin-1β,IL-1β）是一种重要的促炎细胞因子,在中枢神经系统中发挥着广泛的生物学功能。大量研究表明,IL-1β的作用非常复杂,在不同的模型和条件下作用不同,包括神经损伤或者神经毒性作用。电压门控钾通道调节神经元电学性质,也参与多种中枢神经系统的病理学过程。虽然IL-1β和钾通道都在脑损伤和脑疾病过程中发挥重要作用,但目前还很少有它们之间关系的研究报导。文章作者以原代培养的大鼠皮层神经元为材料,使用全细胞膜片钳技术,研究了10 ng/mL的IL-1β在不同处理时间下对皮层神经元电压依赖性钾电流的影响。根据电流的性质,可以将记录到的钾电流分为瞬时外向电流（IA）为主的IA样电流和延迟整流电流（IK）为主的IK样电流两部分,结果显示：IL-1β处理8 h对二者没有作用;处理24 h可使IA样和IK样电流的幅度降低20%左右。以上结果提示IL-1β对大鼠皮层神经元电压门控钾电流具有抑制作用,并且这种抑制可能具有时间依赖性。     

Neurons differentiating from murine neural crest in culture exhibit sensory or sympathetic‐like calcium currents

The trunk neural crest gives rise to peripheral sensory and sympathetic neurons. In culture, neural crest cells can be induced to differentiate into either neuronal phenotype. Few studies have examined the differentiation of physiological properties in cultures of neural crest cells. Using whole‐cell recordings, our study examined the effects of growth factors on high‐voltage‐activated calcium current profiles exhibited by neurons differentiating in culture. We compared these profiles with those exhibited by sensory and sympathetic neurons. Neural crest cells in culture gave rise to neurons with calcium current profiles identical to either sensory or sympathetic neurons, depending on the growth conditions. On average, the calcium current profile for sensory neurons was 23% (L), 51% (N), and 12% (P), while sympathetic neurons had a similar L‐type current (20%), higher N‐type (76%), and lower P‐type (4%). Neural crest cells cultured with human leukemia inhibitory factor plus somite cells produced neurons with a sympathetic‐like calcium current profile (L: 17%, N: 75%, and P: 4%). However, murine leukemia inhibitory factor (L: 25%, N: 52%, and P: 13%) and ciliary neurotrophic factor (L: 18%, N: 49%, and P: 9%) plus somite cells produced neurons with sensory‐like calcium current profiles. These growth conditions did not modify the calcium current profiles of neurons cultured from embryonic and neonatal ganglia. Similarly, murine leukemia inhibitory factor produced a greater percentage of neurons (57%) with sensitivity to capsaicin (sensory phenotype) than human leukemia inhibitory factor (3%). Physiological traits can be a useful tool for the determination of neuronal phenotype in culture where other traits may be less stable. © 1999 John Wiley & Sons, Inc. J Neurobiol 39: 501–514, 1999     

Potassium currents in presynaptic hair cells of Hermissenda.

Apart from their primary function as balance sensors, Hermissenda hair cells are presynaptic neurons involved in the Ca(2+)-dependent neuronal plasticity in postsynaptic B photoreceptors that accompanies classical conditioning. With a view to beginning to understand presynaptic mechanisms of plasticity in the vestibulo-visual system, a locus for conditioning-induced neuronal plasticity, outward currents that may govern the excitability of hair cells were recorded by means of a whole-cell patch-clamp technique. Three K+ currents were characterized: a 4-aminopyridine-sensitive transient outward K+ current (IA), a tetraethyl ammonium-sensitive delayed rectifier K+ current (IK,V), and a Ca(2+)-activated K+ current (IK,Ca). IA activates and decays rapidly; the steady-state activation and inactivation curves of the current reveal a window current close to the apparent resting voltage of the hair cells, suggesting that the current is partially activated at rest. By modulating firing frequency and perhaps damping membrane oscillations, IA may regulate synaptic release at baseline. In contrast, IK,V and IK,Ca have slow onset and exhibit little or no inactivation. These two K+ currents may determine the duration of the repolarization phase of hair-cell action potentials and hence synaptic release via Ca2+ influx through voltage-gated Ca2+ channels. In addition, IK,Ca may be responsible for the afterhyperpolarization of hair cell membrane voltage following prolonged stimulation.     

Voltage-dependent calcium current and the effects of adrenergic modulation in rat aortic smooth muscle cells.

Smooth muscle cells from rat aorta were cultured in defined, serum-free medium and studied using whole-cell patch-clamp techniques. Under conditions designed to isolate currents through Ca channels, step depolarizations produced inward currents which were fast in onset and inactivated rapidly, with little sustained inward current being observed. Both Ni and Cd blocked these currents, with Ni being effective at 50 microM. Removal of external Na or addition of 1 microM tetrodotoxin had no effect. Peak inward currents were attained at about -15 mV, with half-maximal activation at -41 mV using -80 mV holding potentials. The transient inward currents were reduced by depolarized holding potentials, with half-maximal steady-state inactivation at -48 mV. In three of the 98 cells studied, small maintained inward currents were observed with a -40 mV holding potential. The Ca channel antagonist nicardipine (5 microM) blocked the transient inward current while neither of the dihydropyridine Ca channel agonists S(+)202 791 and (-)BAY K 8644 produced a significant augmentation of sustained inward current. At 10 microM, both noradrenaline and adrenaline but not phenylephrine decreased the peak inward current. This inhibition was unaffected by a variety of adrenoceptor antagonists and was also observed when internal solutions having high Ca buffering capacity were used, but was absent when GDP-beta-S instead of GTP was included in the pipette solution. The main conclusions from this study are that under our cell culture conditions, rat aortic smooth muscle cells possess predominantly a transient, low-threshold-activated inward Ca current and that this Ca current is inhibited by certain adrenoceptor agonists but with a quite atypical adrenoceptor antagonist pharmacology.     

P2X2 receptors differentiate placodal vs. neural crest C-fiber phenotypes innervating guinea pig lungs and esophagus

The lungs and esophagus are innervated by sensory neurons with somata in the nodose, jugular, and dorsal root ganglion. These sensory ganglia are derived from embryonic placode (nodose) and neural crest tissues (jugular and dorsal root ganglia; DRG). We addressed the hypothesis that the neuron's embryonic origin (e.g., placode vs. neural crest) plays a greater role in determining particular aspects of its phenotype than the environment in which it innervates (e.g., lungs vs. esophagus). This hypothesis was tested using a combination of extracellular and patch-clamp electrophysiology and single-cell RT-PCR from guinea pig neurons. Nodose, but not jugular C-fibers innervating the lungs and esophagus, responded to alpha,beta-methylene ATP with action potential discharge that was sensitive to the P2X3 (P2X2/3) selective receptor antagonist A-317491. The somata of lung- and esophagus-specific sensory fibers were identified using retrograde tracing with a fluorescent dye. Esophageal- and lung-traced neurons from placodal tissue (nodose neurons) responded similarly to alpha,beta-methylene ATP (30 microM) with a large sustained inward current, whereas in neurons derived from neural crest tissue (jugular and DRG neurons), the same dose of alpha,beta-methylene ATP resulted in only a transient rapidly inactivating current or no detectable current. It has been shown previously that only activation of P2X2/3 heteromeric receptors produce sustained currents, whereas homomeric P2X3 receptor activation produces a rapidly inactivating current. Consistent with this, single-cell RT-PCR analysis revealed that the nodose ganglion neurons innervating the lungs and esophagus expressed mRNA for P2X2 and P2X3 subunits, whereas the vast majority of jugular and dorsal root ganglia innervating these tissues expressed only P2X3 mRNA with little to no P2X2 mRNA expression. We conclude that the responsiveness of C-fibers innervating the lungs and esophagus to ATP and other purinergic agonists is determined more by their embryonic origin than by the environment of the tissue they ultimately innervate.