The role of slow potassium current in phenomena of voltage-dependent action of 4-aminopyridine in amphibian myelinated nerve fibres

The mechanism underlying the voltage-dependent action of 4-aminopyridine (4-AP) is investigated in experiments on amphibian myelinated nerve fibres (Rana ridibunda Pallas) by way of extracellular recording of electrical activity and using activators of potassium current (potassium-free solution and nitric oxide NO) and inhibitors of sodium current (tetrodotoxin). Measurement of action potential (AP) areas was used to evaluate the extent of general membrane depolarization during the activity of nerve fibres. Tetrodotoxin-induced decrease in general membrane depolarization (when the action potential amplitude was reduced by less than 20%) leads to an increase in the duration of depolarizing after-potential (DAP). This supports the dependence of time course of DAP in the presence of 4-AP on ratio of fast and slow potassium channels. In the absence of 4-AP, potassium-free solution and NO increase the potassium current through fast potassium channels (decreasing AP duration, reducing DAP and sometimes producing fast hyperpolarizing after-potential (HAP) after shortened AP), and in the presence of 4-AP these activators increase potassium current through unblocked slow potassium channels (making the development of slow HAP induced by 4-AP more rapid). The increase of slow HAP induced by 4-AP under the influence of potassium-free solution with NO supports the idea that slow HAP is due to activation of slow potassium channels and argues against the notion of removal of block of fast potassium channels. All analyzed phenomena of voltage-dependent action of 4-AP in amphibian myelinated nerve fibers can be accounted for by the activation of slow potassium current produced by membrane depolarization and a decrease of the amount of fast potassium channels involved in the membrane repolarization.     

Changes in Electrogenesis in the Motor Nerve Terminal with Long-Lasting High-Frequency Activation as a Factor of the Control of Transmitter Release

Using the technique of extracellular recording from the region of the neuromuscular junction in the cutaneous-sternal muscle in the frog under conditions of a reduced concentration of Ca²⁺ in the surrounding milieu, we demonstrated that long-lasting (10 min) rhythmic stimulation of the motor nerve with a frequency of 10 sec^{− 1} leads to a gradual increase in the evoked transmitter release. These changes are accompanied by a decrease in the amplitude of electrical responses of the nerve terminal (NT) and by a retardation of its second phase, as well as by a diminution of the third phase. Under conditions of long-lasting (5 min) stimulation with a frequency of 50 sec⁻¹, we observed a two-phase change in the intensity of transmitter release: on the 2nd min, the initial rise was replaced by inhibition. Modifications of the response of the NT with different stimulation frequencies were qualitatively similar, but with a frequency of 10 sec⁻¹ they were clearly expressed. Mathematical simulation of ion currents in the NT demonstrated that voltage-dependent potassium and sodium channels are inactivated in the course of long-lasting high-frequency excitation; the shape of the action potential is modified with changes in the rate of such inactivation. This leads to either an increase or a decrease of the inward calcium current. We conclude that the change in electrogenesis in the NT with long-lasting high-frequency activation of neuromuscular junctions exerts a significant influence on the dynamics of transmitter release. Neirofiziologiya/Neurophysiology, Vol. 37, No. 2, pp. 108–115, March–April, 2005.     

A distributed parameter identification problem in neuronal cable theory models

Dendritic and axonal processes of nerve cells, along with the soma itself, have membranes with spatially distributed densities of ionic channels of various kinds. These ionic channels play a major role in characterizing the types of excitable responses expected of the cell type. These densities are usually represented as constant parameters in neural models because of the difficulty in experimentally estimating them. However, through microelectrode measurements and selective ion staining techniques, it is known that ion channels are non-uniformly spatially distributed. This paper presents a non-optimization approach to recovering a single spatially non-uniform ion density through use of temporal data that can be gotten from recording microelectrode measurements at the ends of a neural fiber segment of interest. The numerical approach is first applied to a linear cable model and a transformed version of the linear model that has closed-form solutions. Then the numerical method is shown to be applicable to non-linear nerve models by showing it can recover the potassium conductance in the Morris-Lecar model for barnacle muscle, and recover the spine density in a continuous dendritic spine model by Baer and Rinzel.     

Local development of action potentials in slow muscle fibres after complete or partial denervation.

Pyriformis muscles of Rana temporaria were completely or partially denervated by cutting the sciatic nerve or some of the small nerve branches entering the muscle. One stimulating and one to three recording microelectrodes were inserted along the fibres in order to compare the electrical activity at these points. In an early period following denervation action potentials of variable size and shape could be observed; these action potentials were often composed of two, sometimes of three or four, components. The size of individual components depended on the position of the recording microelectrode. Individual components could occasionally be triggered separately by adjusting the strength of the stimulating current pulse; propagation of these "all or none" responses was absent. In other fibres one component of the action potential could trigger another one several millimetres apart, thus indicating propagation. Conduction velocities were approximately 0.4 m/s. In partially denervated slow fibres, endplate potentials were confined to one lateral segment of the fibres, while the action potential occupied the denervated part of the membrane. The amplitudes of endplate and action potentials varied inversely with distance. Rough estimates of the length constant of the slow fibre membrane were calculated from the spatial decay of action potentials, endplate potentials and hyperpolarizing electrotonic potentials; mean values obtained were 2.5, 4.8 and 7.7 mm respectively. The results suggest that following denervation Na channels are built into discrete areas of the slow fibre membrane and that this process depends on the amount of denervation in individual fibres.     

How different two almost identical action potentials can be: A model study on cardiac repolarization

Spatial heterogeneity in the properties of ion channels generates spatial dispersion of ventricular repolarization, which is modulated by gap junctional coupling. However, it is possible to simulate conditions in which local differences in excitation properties are electrophysiologically silent and only play a role in pathological states. We use a numerical procedure on the Luo-Rudy phase 1 model of the ventricular action potential (AP1) in order to find a modified set of model parameters which generates an action potential profile (AP2) almost identical to AP1. We show that, although the two waveforms elicited from resting conditions as a single AP are very similar and belong to membranes sharing similar passive electrical properties, the modified membrane generating AP2 is a weaker current source than the one generating AP1, has different sensitivity to up/down-regulation of ion channels and to extracellular potassium, and a different electrical restitution profile. We study electrotonic interaction of AP1- and AP2 - type membranes in cell pairs and in cable conduction, and find differences in source-sink properties which are masked in physiological conditions and become manifest during intercellular uncoupling or partial block of ion channels, leading to unidirectional block and spatial repolarization gradients. We provide contour plot representations that summarize differences and similarities. The present report characterizes an inverse problem in cardiac cells, and strengthen the recently emergent notion that a comprehensive characterization and validation of cell models and their components are necessary in order to correctly understand simulation results at higher levels of complexity.     

Voltage gated ionic channels in rat cultured astrocytes, reactive astrocytes and an astrocyte-oligodendrocyte progenitor cell

Astrocytes (both type 1 and type 2), cultured from the central nervous system of newborn or 7 day old rats show voltage gated sodium and potassium channels that are activated when the membrane is depolarized to greater than -40 mV. The sodium channels in these cells have an h-infinity curve similar to that of nodal membranes but the activation (peak current-voltage) curves are shifted along the voltage axis by about +30 mV. These sodium currents are blocked only by high concentrations of tetrodotoxin. The voltage activated potassium currents in both types of astrocyte show at least two components; an inactivating component that is suppressed at holding potentials of greater than -40 mV and a persistent, non-inactivating current. Several types of single channel currents were observed in outside-out membrane patches from type 2 astrocytes. One type of potassium channel showed inactivation on depolarization and may contribute to the whole-cell inactivating current. In contrast, oligodendrocytes showed no obvious voltage gated membrane channels. The properties of the type 2 astrocyte-oligodendrocyte progenitor cell were investigated in two ways: 1) by examination of cells just beginning to differentiate along the "electrically silent" oligodendrocyte pathway or 2) by recording from progenitor cells cultured for 24 hours in the presence of cycloheximide to block the appearance of new membrane channels. In both cases, voltage gated inward (sodium) and outward (potassium) currents were noted. The outward current response showed both an inactivating and a non-inactivating component. Similar voltage activated inward and outward membrane currents were noted in reactive astrocytes freshly isolated (3-6 hours) from lesioned areas of adult rat brains.(ABSTRACT TRUNCATED AT 250 WORDS)     

BK Channels Reveal Novel Phosphate Sensitivity in SNr Neurons

Whether large conductance Ca²⁺-activated potassium (BK) channels are present in the substantia nigra pars reticulata (SNr) is a matter of debate. Using the patch-clamp technique, we examined the functional expression of BK channels in neurons of the SNr and showed that the channels were activated or inhibited by internal high-energy phosphates (IHEPs) at positive and negative membrane potentials, respectively. SNr neurons showed membrane potential hyperpolarization under glucose-deprivation conditions which was attenuated by paxilline, a specific BK channel blocker. In addition, Fluo-3 fluorescence recording detected an increase in the level of internal free calcium ([Ca²⁺]_i) during ischemic hyperpolarization. These results confirm that BK channels are present in SNr neurons and indicate that their unique IHEP sensitivity is requisite in neuronal ischemic responses. Bearing in mind that the K_ATP channel blocker tolbutamide also attenuated the hyperpolarization, we suggest that BK channels may play a protective role in the basal ganglia by modulating the excitability of SNr neurons along with K_ATP channels under ischemic stresses.     

The effect of acetylcholine on Characeae K+ channels at rest and during action potential generation

The role of acetylcholine (ACh) as a signalling molecule in plants was investigated using a model system of Characeae cells. The effect of ACh on conductance of K⁺ channels in Nitella flexilis cells and on the action potential generation in Nitellopsis obtusa cells after H⁺-ATPase inhibition, where repolarization occurs after the opening of outward rectifying K⁺ channels, was investigated. Voltage-clamp method based on only one electrode impalement was used to evaluate the activity of separate potassium ion transport system at rest. We found that ACh at high concentrations (1 mM and 5 mM) activates K⁺ channels as the main membrane transport system at the resting state involved in electrogenesis of Characeaen membrane potential. We observed that ACh caused an increase in duration of AP repolarization of cells in K⁺ state when plasmalemma electrical characteristics are determined by large conductance K⁺ channels irrespective of whether AP were spontaneous or electrically evoked. These results indicate interference of ACh with electrical cellular signalling pathway in plants.     

Conduction along myelinated and demyelinated nerve fibres with a reorganized axonal membrane during the recovery cycle: model investigations

The changes in the excitability of the reorganized axonal membrane in myelinated and demyelinated nerve fibres as well as the causes conditioning such changes have been investigated by paired stimulation during the first 30 ms of the recovery cycle. The variations of the action potential parameters (amplitude and velocity) are traced also. The simulation of the conduction along the normal fiber is based on the Frankenhaeuser and Huxley (1964) and Goldman and Albus (1968) equations, while the demyelination is considered to be an elongation of the nodes of Ranvier. The axonal membrane reorganization is achieved by means of potassium channel blocking and increase of the sodium-channel permeability. It is shown that potassium channels block decreases membrane excitability for the myelinated and demyelinated fibres in the cases of initial and paired stimulation. With increasing sodium-channel permeability on the background of the blocked potassium channels, the membrane excitability is increased. For the fibres with a reorganized membrane, a supernormality of the membrane excitability is obtained, the latter remaining unrecovered during the 30 ms cycle under investigation. The supernormality of the excitability grows from the demyelinated fibre without reorganized membrane to the demyelinated fibre with reorganized one. For short interstimulus intervals, the second action potential propagates along the fibres with a reduced velocity and a decreased amplitude. No supernormality of the potential parameters (amplitude, velocity) is observed during the cycle up to 30 ms. The membrane properties of the myelinated and demyelinated fibres with blocked potassium channels recover in the interval from 15 to 20 ms depending on whether the sodium channels' increase of the permeability is added on the background of the blocked potassium channel or not. In the recovery cycle, the axonal membrane reorganization leads to an improvement of the conduction along most severely demyelinated fibres.     

The orientation and molecular movement of a k(+) channel voltage-sensing domain

Voltage-gated channels operate through the action of a voltage-sensing domain (membrane segments S1-S4) that controls the conformation of gates located in the pore domain (membrane segments S5-S6). Recent structural studies on the bacterial K(v)AP potassium channel have led to a new model of voltage sensing in which S4 lies in the lipid at the channel periphery and moves through the membrane as a unit with a portion of S3. Here we describe accessibility probing and disulfide scanning experiments aimed at determining how well the K(v)AP model describes the Drosophila Shaker potassium channel. We find that the S1-S3 helices have one end that is externally exposed, S3 does not undergo a transmembrane motion, and S4 lies in close apposition to the pore domain in the resting and activated state.     

Changes in membrane potential and resistance caused by transient increase of potassium conductance in the unicellular green alga Eremosphaera viridis

The electrophysiological membrane parameters of the unicellular green alga Eremosphaera viridis were determined using an improved computer-supported single-microelectrode technique. These cells developed an average membrane potential of-150 mV in the light and a specific resistance of 1 Ω m² with an external potassium concentration of 1.1 mM and pH 5.5. In the dark, many cells showed a less polarized potential of 30–40 mV and a smaller membrane resistance. At potassium concentrations in the external medium higher than 1 mM, the membrane potential strongly depends on the external potassium content apart from a small electrogenic component. At concentrations lower than 1 mM K⁺, a dependence of the membrane potential upon external potassium concentrations could not be verified. Inserting the internal ion activities in the Goldmann equation shows that, in this range, the proton conductance seems to be predominant over the potassium conductance. Transient changes in the membrane potential and in the membrane resistance were observed after switching off the light, after addition of 3-(3′,4′-dichlorophenyl)-1,1-dimethylurea or N,N′-dicyclohexylcarbodiimide, after a sudden decrease in temperature, and after current pulses. These changes resemble the action potentials (AP) found in other plant cells (Chara, Acetabularia). On average, the AP has a delay period of 5.1 s and a duration of 43.8 s showing a sudden decrease and a slower regeneration. The voltage peak during an AP followed exactly the Nernst potential of potassium over a range of external potassium concentrations from 5 μM to 0.2 M. This is true for depolarization or hyperpolarization, depending on the external K⁺-concentration. Tetraethylammonium-hydrogensulphate, a rather specific inhibitor of K⁺ channels in nervous cells, suppressed the AP. The correlation of the appearance of the AP with a short-term opening of potassium channels in the membrane of Eremosphaera is discussed.     

Expression of voltage dependent potassium currents in freshly dissociated rat articular chondrocytes.

The electrophysiological properties of voltage dependent potassium channels from freshly dissociated rat articular chondrocytes were studied. The resting membrane potential (-42.7+/-2.0 mV) was significantly depolarized by increasing concentrations of external potassium. No change was observed when external chloride concentration was varied. Addition of TEA, 4AP, alpha-Dendrotoxin and charybdotoxin depolarized resting membrane potential. Whole cell patch clamp studies revealed the presence of outwardly rectifying currents whose kinetic and pharmacological properties suggest the expression of voltage dependent potassium channels. Two kinds of currents were observed under the same experimental conditions. The first one, most frequently observed (80%), starts activating near -50 mV, with V(1/2)=-18 mV, G(max)=0.30 pS/pF. The second kind was observed in only 10% of cases; It activates near -40 mV, with(1/2)=+28.35 mV, G(max)=0.28 pS/pF pA/pF and does not inactivates. Inactivating currents were significantly inhibited by TEA (IC(50)=1.45 mM), 4AP (IC(50)=0.64 mM), CTX (IC(50) = 10 nM), alpha-Dendrotoxin (IC(50) < 100 nM) and Margatoxin (IC(50)=28.5 nM). These results show that rat chondrocytes express voltage dependent potassium currents and suggest a role of voltage-dependent potassium channels in regulating membrane potential of rat chondrocytes.     

Calcium oscillations induced by gambierol in cerebellar granule cells

Gambierol is a marine polyether ladder toxin derived from the dinoflagellate Gambierdiscus toxicus. To date, gambierol has been reported to act either as a partial agonist or as an antagonist of sodium channels or as a blocker of voltage‐dependent potassium channels. In this work, we examined the cellular effect of gambierol on cytosolic calcium concentration, membrane potential and sodium and potassium membrane currents in primary cultures of cerebellar granule cells. We found that at concentrations ranging from 0.1 to 30 µM, gambierol‐evoked [Ca²⁺]c oscillations that were dependent on the presence of extracellular calcium, irreversible and highly synchronous. Gambierol‐evoked [Ca²⁺]c oscillations were completely eliminated by the NMDA receptor antagonist APV and by riluzole and delayed by CNQX. In addition, the K⁺ channel blocker 4‐aminopyridine (4‐AP)‐evoked cytosolic calcium oscillations in this neuronal system that were blocked by APV and delayed in the presence of CNQX. Electrophysiological recordings indicated that gambierol caused membrane potential oscillations, decreased inward sodium current amplitude and decreased also outward IA and IK current amplitude. The results presented here point to a common mechanism of action for gambierol and 4‐AP and indicate that gambierol‐induced oscillations in cerebellar neurons are most likely secondary to a blocking action of the toxin on voltage‐dependent potassium channels and hyperpolarization of sodium current activation. J. Cell. Biochem. 110: 497–508, 2010. © 2010 Wiley‐Liss, Inc.     

Potassium channel gating in adhesion: from an oocyte–silicon to a neuron–astrocyte adhesion contact

In a neuron–astrocyte adhesion contact the ionic current due to the opening of voltage-dependent potassium channels has to flow along a narrow intercellular cleft, generating there an extracellular voltage. This voltage might be large enough to affect significantly the dependence of channel gating from the intracellular voltage. In order to test this hypothesis, we considered a Xenopus oocyte expressing voltage-dependent potassium channels adhering to a layer of silicon oxide as a simplified model of cell–cell adhesion; here the cell membrane and silicon oxide are separated by a narrow cleft and form a junction of circular shape. We measured directly the extracellular voltage along the diameter of the cleft and investigated its effect on channel gating using a linear array of field effect transistors integrated in the silicon substrate. On this experimental basis we demonstrated that the voltage dependence of potassium channels is strongly affected by adhesion, as can be predicted using a model of a two-dimensional cable and electrodiffusion theory. Computations based on the model showed that along a neuron–astrocyte adhesion contact the opening of voltage-dependent Kv2.1 potassium channels is significantly reduced with respect to identical channels facing an open extracellular space.     

海马脑片缺氧早期腺苷的作用及其机制研究

本实验采用海马脑片细胞外记录技术,观察了缺氧早期突触功能可逆性抑制中腺苷的作用并初步探讨其作用机制。结果发现：海马脑片缺氧早期突触功能出现可逆性抑制,与外源施加高浓度腺苷反应类同。腺苷Ａ１受体拮抗剂ＣＰＴ以及Ｋ＋通道阻断剂４－ＡＰ可阻断这种抑制作用;而ＴＥＡ以及ＡＴＰ敏感Ｋ＋通道阻断剂ｇｌｉｐｉｚｉｄｅ均未见显著效应。结果提示：缺氧早期突触功能可逆性抑制与内源性腺苷大量释放有关,腺苷通过作用其Ａ１受体,激活４－ＡＰ敏感Ｋ＋通道,从而抑制突触传递,显示其抗缺氧作用。ＡＴＰ敏感性Ｋ＋通道可能不参于这个过程。     

The presence of voltage-gated sodium, potassium and chloride channels in rat cultured astrocytes

Patch-clamp recording from the plasmalemma of rat cultured astrocytes reveals the presence of both voltage-dependent sodium and voltage-dependent potassium conductances. These conductances are similar but not identical to the corresponding conductances in the axolemma. Whereas the h infinity relation of the sodium channels has the same voltage dependence as in the nodal axolemma, the peak current-voltage relation is shifted by about 30 mV along the voltage axis in the depolarizing direction. It is speculated that the glial cells synthesize sodium and potassium channels for later insertion into the axolemma of neighbouring axons. The astrocytes also express a plasmalemmal voltage-dependent anion conductance that is turned on at about -40 mV (that is, near the resting potential of the cultured astrocytes). The channels involved are large enough to be just permeable to glutamate but not to ascorbate. It is suggested that the conductance of this channel for chloride plays a physiological role in the spatial buffering of potassium by glial cells.     

A mathematical model of a myelinated nerve fiber for analysis of impulse conduction mechanisms during the relative refractory period

It was shown by means of a mathematical model of a myelinated nerve fiber (Frankenhaeuser — Huxley) that an increase in threshold and decrease in the amplitude of the action potential (AP) during the relative refractory period are due mainly to sodium inactivation. The contribution of increased potassium permeability to these changes is small, for the chief component of the outgoing ionic current in the node of Ranvier is not the potassium current, but the leak current. Given the ratio between these currents the increase in threshold and graduation of the action potential in the node membrane are less marked than in the membrane of the squid giant axon. At the beginning of the relative refractory period the AP evoked by strong stimulation is conducted only to the next node. Later in the refractory period impulses are conducted incrementally, and the threshold for the spreading impulse is higher than the threshold for spike excitation in the stimulated node. Delay in impulse conduction between refractory nodes leads to the formation of a retrograde depolarization wave. The reasons for differences in the mechanisms of impulse conduction along unmyelinated and myelinated refractory fibers are discussed.Vishnevskii Institute of Surgery, Academy of Medical Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 4, No. 2, pp. 201–207, March–April, 1972.     

Effects of pyrocatechol on neuromuscular transmission

Effects of pyrocatechol on neuromuscular transmission were studied both in the frog pectoral-cutaneous muscle and in the mouse phrenic-diaphragmatic preparation by means of extracellular microelectrode recording of synaptic signals. Pyrocatechol applied in a concentration of 0.05 mM increased the frequency of miniature end-plate currents (MEPC) and the amplitude of end-plate current (EPC) by increasing its quantum content. Pyrocatechol also increased the duration of presynaptic response. When voltage-dependent potassium channels had been blocked, pyrocatechol affected neither the EPC quantum content nor the duration of presynaptic response. It is suggested that the pyrocatechol-induced enhancement of transmitter release results from modulatory effects of pyrocatechol on voltage-dependent potassium current in the membrane of a nerve terminal.Neirofiziologiya/Neurophysiology, Vol. 25, No. 6, pp. 405–408, November–December, 1993.     

基于随机森林法的山核桃林地土壤速效养分含量空间分布特征研究

土壤氮磷钾是土壤肥力管理的重要元素,是植物生长的必要养分元素。对土壤氮磷钾的空间分布进行特征解译,可为精准管理临安山核桃产区林地土壤肥力,促进山核桃林产业可持续发展提供理论依据。研究以临安山核桃主产区为研究区域,利用随机森林（RF）、普通克里格（OK）和Shapley加性解释（SHAP）方法,结合地形因子、气候因子、土壤因子、遥感因子等环境变量,对山核桃林地土壤碱解氮（AN）、有效磷（AP）、速效钾（AK）的空间分布特征进行分析。研究结果表明：相比于OK模型,基于环境协变量所构建的RF模型对AN、AP和AK含量空间分布预测表现最佳,R²分别为0.68、0.60和0.64,均方根误差（RMSE）分别为20.005、10.287和22.426,平均绝对误差（MAE）分别为15.425、7.709 和21.628。RF模型SHAP分析显示,AN和AK含量分布主要受土壤有机质（SOM）的影响,并且SOM与AN和AK存在正相关性;AP主要受pH的影响,其次为色调指数,AP与pH和色调指数均具有负相关性;AK和AP同时受到海拔和坡向的影响。两种模型预测的氮磷钾空间分布趋势总体相似,不同速效养分存在明显的空间异质性。碱解氮高值区域主要分布于研究区东部;有效磷高值区域主要分布于研究区西部,但分散度高;速效钾高值区域则主要分布于研究区中部。总体而言,基于随机森林模型可以高精度模拟山核桃林地土壤氮磷钾含量空间分布特征,并依据主要环境协变量对土壤氮磷钾的影响关系,提出相应改良措施。在有效磷含量低值区域可以施用石灰来缓解土壤酸化,同时补追磷肥;碱解氮含量高值区域可以合理减少氮肥施用;速效钾含量低值区域合理施加钾肥;对于海拔较高及迎风坡多降雨的区域,可以构建林下高效水土保持植被,减轻水土流失;在林地施用有机肥料,改善土壤理化性质,增加土壤养分含量。     

Neurotransmission in the medulla mediating insular cortical and lateral hypothalamic sympathetic responses

Previous evidence has shown sympathetic nerve responses to insular cortical (IC) stimulation are mediated by synapses within the lateral hypothalamic area (LHA) and ventrolateral medulla (VLM). The present study was aimed at determining the neurotransmitter(s) and receptor(s) involved at the synapse in the VLM. Twenty male Wistar rats were instrumented for renal nerve, arterial pressure, and heart rate recording. The IC or the LHA was stimulated with a bipolar electrode (200-1000 microA; 2 ms; 0.8 Hz) to elicit sympathetic nerve responses. Antagonists were then pressure-injected into the VLM (300 nL). Bilateral and unilateral kynurenate (25 mM) resulted in 100% block of IC-and LHA-stimulated sympathetic nerve responses. Bilateral injection of the non-NMDA (N-methyl-D-aspartate) receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 200 microM) also resulted in up to 100% block of IC and LHA sympathetic responses. In addition, unilateral injections of CNQX were made in two animals, resulting in 100 and 83% block of LHA sympathetic responses. Bilateral injection of the NMDA receptor antagonist DL-2-amino-5-phosphonopentanoic acid (AP5; 200 microM) did not affect the response to IC or LHA stimulation. Kynurenate, CNQX, and AP5 all resulted in an elevation of baseline sympathetic nerve activity and a pressor response. Kynurenate resulted in a 263+/-79% increase in baseline activity, while CNQX and AP5 resulted in 83+/-19% and 91+/-21% increases. respectively. Bilateral injections of antagonists for GABA(A) (bicuculline; 0.1 microM), acetylcholine (atropine; 0.1 microM) and catecholaminergic alpha and beta receptors (phentolamine and propranolol: 0.1 microM) had no effect on LHA sympathetic responses. Thus, sympathetic responses originating in the IC and LHA are mediated by a non-NMDA receptors in the VLM, which are likely AMPA receptors.