Effects of GABA, THIP, and potassium on excitability of myelinated axons of isolated amphibian spinal roots

The effects of direct applications of GABA (gamma-aminobutyric acid) and the GABAA agonist, THIP (4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol) on the excitability of myelinated axons of individual dorsal and ventral spinal roots (lumbar VI and (or) VII) of the isolated bullfrog peripheral nerve are reported. Increases evoked by the GABA agonists (0.01-10 mM) in the amplitude of half-maximal A-fiber compound action potentials indicate the presence of depolarizing responses with apparently greater localization to the dorsal roots, and a sensitivity to GABA twofold greater than that for THIP. The changes evoked by GABA and THIP, as well as potassium have components that closely resemble those of sensory and motor fibers in the more distal, desheathed nerve bundle but are smaller and delayed, differences attributable to a closely attached root sheath that acts as a diffusion barrier. These results confirm the likely existence of GABAA receptors on both dorsal and ventral spinal roots.     

Effects of barium on isolated frog spinal cord

The effects of Ba2+ were studied in vitro on the isolated frog spinal cord. Ba2+ (25 microM-5 mM) caused a concentration-dependent depolarization of ventral (VR) and dorsal (DR) roots. TTX and Mg2+ substantially reduced the depolarization suggesting that interneuronal effects were involved. Ba2+ (25-500 microM) markedly increased the frequency and duration of spontaneous VR and DR potentials and substantially enhanced the duration (and frequently the amplitude) of VR and DR potentials evoked by DR stimulation. Higher concentrations of Ba2+ (1-5 mM) reduced both spontaneous and evoked potentials. Ba2+ (25-500 microM) enhanced the amount of K+ released by a DR volley and by application of L-glutamate and L-aspartate. The cation reduced VR and DR root depolarizations produced by elevated [K+]0. VR potentials induced by L-glutamate, L-aspartate, GABA and glycine and DR depolarizations caused by GABA were reduced by Ba2+. These results show that Ba2+ has complex actions on reflex transmission, interneuronal activity, the postsynaptic actions of excitatory and inhibitory amino acids and the evoked release of K+.     

Modulating action of barium and other ions on the sensory activity of frog labyrinth

The effect of Ba2+, TEA, 4-AP and CoCl2 on the EPSP and spike discharges recorded from single fibres of the posterior nerve in the isolated frog labyrinth has been investigated. In Ca-free solution Ba2+ preserved, at low concentration (0.3 mM), the resting activity and at higher levels (up to 6 mM) it resulted in a pronounced facilitation of the EPSP and spike discharges. Facilitation increased on increasing Ba2+ concentration up to 4-5 mM and it was more evident in those units exhibiting a low resting spike firing. The effect of Ba2+ (1 mM) was completely antagonized by 10 mM Ca2+ X CoCl2 (3 mM) suppressed the resting rate at the normal external Ca2+ concentration; the Co2+ block was partially relieved by 1.8 mM Ba2+ X TEA (20 mM) evoked a clear-cut increase in the EPSP and spike discharges which, however, was less consistent than that produced by Ba2+. By comparing the effect of TEA on the spike frequency with that obtained at different Ba2+ levels, the Ba2+ capacity to carry the Ca2+ current was dissected. Such an effect is dose-dependent and it is more evident in low-frequency units. Conversely, 4-AP did not affect the resting discharge frequency. These results indicate that either the Ca2+ or the Ba2+ current sustain the transmitter release at the cyto-neural junction. The effect of TEA suggests that the Ca2+-dependent K+ current may play an important role in supporting the neurosecretory process by controlling the membrane potential of the hair cells.     

Excitotoxicity, reflex responses, and evoked changes in extracellular potassium in the frog spinal cord

1. The effects of the excitatory amino acid agonists kainate (KA), quisqualate (QUIS), and N-methyl-D-aspartate (NMDA) were studied in vitro on the hemisected frog spinal cord. 2. Prolonged (1.0 hr) application of excitatory amino acid agonists (KA, 50 or 300 microM; QUIS, 30 microM; NMDA, 300 microM) significantly reduced the ventral root potentials (VRPs) and [K+]0 evoked by a dorsal root tetanus (10 sec, 25 Hz), by brief (10 sec) applications of the same agonists (KA, 30 microM; QUIS, 30 microM; NMDA, 300 microM), and by GABA (10 sec, 1.0 mM). 3. The effect was essentially irreversible and persisted despite 2-4 hr of washing. 4. Excitatory amino acid antagonists (APV, 30 microM and kynurenate, 2 mM) blocked the neurotoxic effects of the excitatory agonists NMDA and KA respectively, an observation which indicates the observed effects of the agonists require the activation of specific excitatory receptors. 5. TTX did not alter the neurotoxic effects of KA suggesting that interneuronal firing does not contribute to the observed changes. 6. Addition of high K+ did not duplicate the effect of prolonged excitatory amino acid agonist exposure, an indication that elevation of K+ does not cause the decreased responses. 7. Light microscopy did not provide any evidence of gross tissue damage. 8. The parallel reduction of postsynaptic responses and delta [K+]0 support the idea that elevation of extracellular [K+] by afferent stimuli results from interneuronal activity.     

A Fast Transient Outward Monovalent Current in Rat Saphenous Myocytes Passing Through Ca2+ Channels

Transient outward currents in rat saphenous arterial myocytes were studied using the perforated configuration of the patch-clamp method. When myocytes were bathed in a Na-gluconate solution containing TEA to block large-conductance Ca2+-activated K+ (BK) currents, depolarizing pulses positive to +20 mV from a holding potential of -100 mV induced fast transient outward currents. The activation and inactivation time constants of the current were voltage dependent, and at +40 mV were 3.6 +/- 0.8 ms and 23.9 +/- 6.4 ms (n = 4), respectively. The steady-state inactivation of the transient outward current was steeply voltage dependent (z = 1.7), with 50% of the current inactivated at -55 mV. The current was insensitive to the A-type K+ channel blocker 4-AP (1-5 mM), and was modulated by external Ca, decreasing to approximately 0.85 of control values upon raising Ca2+ from 1 to 10 mM, and increasing approximately 3-fold upon lowering it to 0.1 mM. Transient outward currents were also recorded following replacement of internal K+ with either Na+ or Cs+, raising the possibility that the current was carried by monovalent ions passing through voltage-gated Ca2+ channels. This hypothesis was supported by the finding that the transient outward current had the same inactivation rate as the inward Ba2+ current, and that both currents were effectively blocked by the L-type Ca2+ channel blocker, nifedipine and enhanced by the agonist BAYK8644.     

Characterization of the Participation of Sodium Channels on the Rise in Na+ Induced by 4-Aminopyridine (4-AP) in Synaptosomes

The participation of voltage-sensitive Na+ channels (VSSC) on the changes on internal (i) Na+, K+, Ca2+, and on DA, Glu, and GABA release caused by different concentrations of 4-AP was investigated in striatum synaptosomes. TTX, which abolished the increase in Na(i) (as determined with SBFI), induced by 0.1 mM 4-AP only inhibited by 30% the rise in Na(i) induced by 1 mM 4-AP. One millimolar 4-AP markedly decreased the fluorescence of the K+ indicator dye PBFI but 0.1 mM 4-AP did not. Like 1 mM 4-AP, ouabain decreased PBFI fluorescence and increased a considerable fraction of Na(i) in a TTX-insensitive manner. In contrast with the different TTX sensitivity of the rise in Na(i) induced by 0.1 and 1 mM 4-AP, the rise in Ca(i) (as determined with fura-2) induced by the two concentrations of 4-AP was markedly inhibited by TTX, as well as by omega-agatoxin in combination with omega-conotoxin GVIA, indicating that only the TTX-sensitive fraction of the rise in Na(i) induced by 4-AP is linked with the activation of presynaptic Ca2+ channels. It is concluded that the TTX-sensitive fraction of neurotransmitter release evoked by 4-AP is released by exocytosis, and the TTX insensitive fraction involves reversal of the neurotransmitters transporters. This contrasts with the exocytosis evoked by high K+ that is unchanged by TTX and with the neurotransmitter-transporter-mediated release evoked by veratridine, which is highly TTX sensitive and does not require activation of Ca2+ channels.     

Low-affinity Na+ uptake in the halophyte Suaeda maritima

Na(+) uptake by plant roots has largely been explored using species that accumulate little Na(+) into their shoots. By way of contrast, the halophyte Suaeda maritima accumulates, without injury, concentrations of the order of 400 mM NaCl in its leaves. Here we report that cAMP and Ca(2+) (blockers of nonselective cation channels) and Li(+) (a competitive inhibitor of Na(+) uptake) did not have any significant effect on the uptake of Na(+) by the halophyte S. maritima when plants were in 25 or 150 mM NaCl (150 mM NaCl is near optimal for growth). However, the inhibitors of K(+) channels, TEA(+) (10 mM), Cs(+) (3 mM), and Ba(2+) (5 mM), significantly reduced the net uptake of Na(+) from 150 mM NaCl over 48 h, by 54%, 24%, and 29%, respectively. TEA(+) (10 mM), Cs(+) (3 mM), and Ba(2+) (1 mm) also significantly reduced (22)Na(+) influx (measured over 2 min in 150 mM external NaCl) by 47%, 30%, and 31%, respectively. In contrast to the situation in 150 mm NaCl, neither TEA(+) (1-10 mM) nor Cs(+) (0.5-10 mM) significantly reduced net Na(+) uptake or (22)Na(+) influx in 25 mM NaCl. Ba(2+) (at 5 mm) did significantly decrease net Na(+) uptake (by 47%) and (22)Na(+) influx (by 36% with 1 mM Ba(2+)) in 25 mM NaCl. K(+) (10 or 50 mM) had no effect on (22)Na(+) influx at concentrations below 75 mM NaCl, but the influx of (22)Na(+) was inhibited by 50 mM K(+) when the external concentration of NaCl was above 75 mM. The data suggest that neither nonselective cation channels nor a low-affinity cation transporter are major pathways for Na(+) entry into root cells. We propose that two distinct low-affinity Na(+) uptake pathways exist in S. maritima: Pathway 1 is insensitive to TEA(+) or Cs(+), but sensitive to Ba(2+) and mediates Na(+) uptake under low salinities (25 mM NaCl); pathway 2 is sensitive to TEA(+), Cs(+), and Ba(2+) and mediates Na(+) uptake under higher external salt concentrations (150 mM NaCl). Pathway 1 might be mediated by a high-affinity K transporter-type transporter and pathway 2 by an AKT1-type channel.     

Pain and neurotransmitters

1. To study physiological roles of substance P (SP), gamma-aminobutyric acid (GABA), enkephalins and other endogenous substances, we developed several kinds of isolated spinal cord preparations of newborn rats. 2. In these preparations, various slow responses of spinal neurons evoked by stimulation of primary afferent C fibers were depressed by a tachykinin antagonist, spantide. These results together with many other lines of evidence suggest that SP and neurokinin A serve as pain transmitters in a subpopulation of primary afferent C fibers. 3. Some C-fiber responses in various isolated spinal cord preparations were depressed by GABA, muscimol, and opioid peptides. In contrast, bicuculline (GABA antagonist) and naloxone (opioid antagonist) potentiated the "tail pinch potential," i.e., a nociceptive response of the ventral root evoked by pinch stimulation of the tail in isolated spinal cord-tail preparation of the newborn rat. The latter results support the hypothesis that some primary afferents activate inhibitory spinal interneurons which release GABA and enkephalins as transmitters to modulate pain inputs.     

Neuronal background two-P-domain potassium channels: molecular and functional aspects

Background K+ conductances are a major determinant of membrane resting potential and input resistance, two key components of neuronal excitability. Background channels have been cloned and form a K+ channel family structurally different from Kv, KCa and Kir channels. These channels with 2P domains (K2P channels) are voltage- and time-independent. They are relatively insensitive to classical potassium channels blockers such as TEA, 4-AP, Ba2+ and Cs+. TASK and TREK subunits are widely expressed in the nervous system. Open at rest, these channels mainly contribute to the resting potential of somatic motoneurons, brainstem respiratory and chemoreceptor neurones, and cerebellar granule cells. K2P channels are regulated by numerous physical and chemical stimuli including extracellular and intracellular pH, temperature, hypoxia, pressure, bioactive lipids, and neurotransmitters. The regulation of these background K+ channels profoundly alters the neuronal excitability. For example, in Aplysia, regulation of a background potassium conductance by neurotransmitters is involved in synaptic modulation, a simple and primitive form of learning. The recent discovery that clinical compounds such as volatile anaesthetics and other neuroprotective agents including riluzole and unsaturated fatty acids activate K2P channels suggest that neuronal background K+ channels are attractive targets for the development of new drugs.     

Antagonist actions of bicuculline methiodide and picrotoxin on extrasynaptic gamma-aminobutyric acid receptors

The effects of picrotoxin and bicuculline methiodide to block depolarizing responses of extrasynaptic receptors for gamma-aminobutyric acid (GABA) are compared using excitability testing of myelinated axons in amphibian peripheral nerve. The actions of the antagonists appear both complex and dissimilar. Picrotoxin (10-1000 microM) produces large reversible depressions of the maximal response to GABA (0.01-10mM) and increases the EC50 from 0.33 to 12.6 mM. With high concentrations of agonist and antagonist an insensitive component is apparent. The action of picrotoxin is not classically noncompetitive: it may represent a mixed antagonism (competitive and noncompetitive) or a noncompetitive one, masked by the presence of receptor reserve and (or) secondary depolarizing influences (e.g., GABA-evoked [K+] o accumulation). Bicuculline methiodide (10-200 microM) shifts the GABA concentration-response curve to the right; maximal responses persist and are even enhanced. The impression that bicuculline methiodide has a competitive action is supported by analysis of its inhibition of responses to low concentrations of the agonist. It is suggested that the enhancement of GABA responses by bicuculline methiodide and their apparent resistance to block by picrotoxin may be due to a common secondary effect of the antagonists such as a decrease in membrane conductance to K+ and (or) block of transmitter uptake.     

Monoaminergic Modulation of Spinal Viscero-Sympathetic Function in the Neonatal Mouse Thoracic Spinal Cord

Descending serotonergic, noradrenergic, and dopaminergic systems project diffusely to sensory, motor and autonomic spinal cord regions. Using neonatal mice, this study examined monoaminergic modulation of visceral sensory input and sympathetic preganglionic output. Whole-cell recordings from sympathetic preganglionic neurons (SPNs) in spinal cord slice demonstrated that serotonin, noradrenaline, and dopamine modulated SPN excitability. Serotonin depolarized all, while noradrenaline and dopamine depolarized most SPNs. Serotonin and noradrenaline also increased SPN current-evoked firing frequency, while both increases and decreases were seen with dopamine. In an in vitro thoracolumbar spinal cord/sympathetic chain preparation, stimulation of splanchnic nerve visceral afferents evoked reflexes and subthreshold population synaptic potentials in thoracic ventral roots that were dose-dependently depressed by the monoamines. Visceral afferent stimulation also evoked bicuculline-sensitive dorsal root potentials thought to reflect presynaptic inhibition via primary afferent depolarization. These dorsal root potentials were likewise dose-dependently depressed by the monoamines. Concomitant monoaminergic depression of population afferent synaptic transmission recorded as dorsal horn field potentials was also seen. Collectively, serotonin, norepinephrine and dopamine were shown to exert broad and comparable modulatory regulation of viscero-sympathetic function. The general facilitation of SPN efferent excitability with simultaneous depression of visceral afferent-evoked motor output suggests that descending monoaminergic systems reconfigure spinal cord autonomic function away from visceral sensory influence. Coincident monoaminergic reductions in dorsal horn responses support a multifaceted modulatory shift in the encoding of spinal visceral afferent activity. Similar monoamine-induced changes have been observed for somatic sensorimotor function, suggesting an integrative modulatory response on spinal autonomic and somatic function.     

Pharmacological characterization of the serotonin-sensitive potassium channel of Aplysia sensory neurons

The effects of a variety of K+ channel blockers on current flow through single serotonin-sensitive K+ channels (the S channels) of Aplysia sensory neurons were studied using the patch-clamp technique. Tetraethylammonium (TEA), 4-aminopyridine (4-AP), and Co2+ and Ba2+ were first applied to the external membrane surface using cell-free outside-out patches. At concentrations up to 10 mM, these agents had little or no effect on single S-channel currents. At higher concentrations, external TEA acted as a fast open-channel blocker, reducing the single-channel current amplitude according to a simple one-to-one binding scheme with an apparent Kd of 90 mM. Blockage by external TEA is voltage independent. Internal TEA also acts as an open-channel blocker, with an apparent Kd of approximately 40 mM and a relatively weak voltage dependence, corresponding to an apparent electrical distance to the internal TEA-binding site of 0.1. Both internal and external TEA block the open channel selectively, with an affinity that is 10-100-fold greater than the affinity for the closed channel. Internal Ba2+ acts as a slow channel blocker, producing long closures of the channel, and binding with an apparent Kd of approximately 25-30 microM. These results show that single S-channel currents share a similar pharmacological profile with the macroscopic S current previously characterized with voltage clamp. On the basis of these results, a structural model for S-channel opening is proposed.     

Hyperpolarization contributes to vascular hyporeactivity in rats with lipopolysaccharide-induced endotoxic shock

We have examined the role of membrane hyperpolarization in mediating vascular hyporeactivity induced by bacterial lipopolysaccharide (LPS) in endothelial-denuded strips of rat thoracic aorta ex vivo. The injection of rats with LPS caused a significant fall of blood pressure and a severe vascular hyporeactivity to norepinephrine. The membrane potential recording showed that endotoxemia caused a hyperpolarization when compared to the control. This hyperpolarization was fully restored by methylene blue (MB; 10 microM) and partially reversed by Nomega-nitro-L-arginine methyl ester (L-NAME; 0.3 mM), 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; 1 microM), tetraethylammonium (TEA; 10 mM), charybdotoxin (CTX; 0.1 microM), or glibenclamide (GB; 10 microM), however, this hyperpolarization was not significantly affected by apamin (0.1 microM), 4-aminopyridine (4-AP; 1 mM), or Ba2+ (50 microM). In addition, the basal tension of the tissues obtained from endotoxemic rats was enhanced by the following order: MB > or = ODQ > TEA > or = L-NAME > or = CTX > GB; whereas apamin, 4-AP or Ba2+ had no significant effects on these tissues. In contrast, none of these inhibitors had significant effects on the membrane potential or the basal tension in control tissues. Our electrophysiological results further confirmed previous studies showing that in addition to nitric oxide, the large conductance Ca2+-activated K+-channels and ATP-sensitive K+-channels are, most likely, responsible for endotoxin-mediated hyporeactivity to vasoconstrictor agents in vascular smooth muscle.     

体外培养新生大鼠大脑皮层星形神经元钾通道的特性及膜外钾离子浓度对其影响

用膜片钳技术中的细胞贴附方式和内面向外方式,首次在新生大鼠大脑皮层星形神经元胞体膜上记录到一类电压依赖性钾通道。此通道可被２０ｍｍｏｌ／Ｌ ＴＥＡ,５ｍｍｏｌ／Ｌ Ｂａ＾２＋,１４０ｍｍｏｌ／Ｌ Ｃｓ＾＋阻断,不受２０ｍｍｏｌ／Ｌ４－ＡＰ影响,其激活不依赖Ｃａ＾２＋。膜外钾离子浓度对通道的特性有显著的影响,逆转电位随Ｋ＾＋的增大而增大,并表现出一定的饱和现象,两者的对数呈线性关系;同一驱动电位下,     

Extracellular potassium rapidly inhibits axonal transport of particles in cultured mouse dorsal root ganglion neurites

Changes in extracellular potassium concentration ([K+]o) modulate a variety of neuronal functions. However, whether axonal transport, which conveys materials to the appropriate destination for morphogenesis and other neuronal functions, depends on the extracellular K+ environment remains unclear. We therefore examined the effects of changes in [K+]o on axonal transport of particles visualized by video-enhanced microscopy in cultured mouse dorsal root gan-glion neurites. Increases in [K+]o (delta[K+]o > or = 2.5 mM) from control concentration (5 mM) inhibited both anterograde and retrograde axonal transport within a few minutes in a concentration-dependent manner. Conversely, removal of extracellular K+ induced the rapid facilitation of transport in both directions. These inhibitory and facilitatory responses were completely blocked by the K+ channel blocker tetraethylammonium (TEA), suggesting that the effect of changes in [K+]o involves the TEA-sensitive K+ channels. Increases in [K+]o provoked membrane depolarization in the absence and presence of TEA. Another depolarizing agent, veratridine, did not produce an effect on axonal transport. These results suggest that the extracellular K+-mediated inhibition of axonal transport does not depend on membrane depolarization. The inhibitory effect of increasing [K+]o on axonal transport was retained in calcium (Ca2+)-free extracellular medium, indicating that the inhibitory effect of extracellular K+ does not result from Ca2+ influx through voltage-dependent Ca2+ channels. In chloride (CI-)-free medium, increasing [K+]o failed to inhibit axonal transport, implying that the extracellular K+-mediated inhibition of axonal transport may be due to an increase in intracellular Cl- concentration associated with increases in the net inward movement of K+ and CI- across the membrane. Our results suggest that the extracellular K+ environment is involved in the rapid modulation of axonal transport of particles in dorsal root ganglion neurites.     

Comparison of changes evoked by GABA (gamma-aminobutyric acid) and anoxia in [K+]o, [Cl-]o, and [Na+]o in stratum pyramidale and stratum radiatum of the guinea pig hippocampus

Ion-selective microelectrode recordings were made to assess a possible contribution of extracellular gamma-aminobutyric acid (GABA) accumulation to early responses evoked in the brain by anoxia and ischemia. Changes evoked by GABA or N2 in [K+]o, [Cl-]o, [Na+]o, and [TMA+]o were recorded in the cell body and dendritic regions of the stratum pyramidale (SP) and stratum radiatum (SR), respectively, of pyramidal neurons in CA1 of guinea pig hippocampal slices. Bath application of GABA (1-10 mM) for approximately 5 min evoked changes in [K+]o and [Cl-]o with respective EC50 levels of 3.8 and 4.1 mM in SP, and 4.7 and 5.6 mM in SR. In SP 5 mM GABA reversibly increased [K+]o and [Cl-]o and decreased [Na+]o; replacement of 95% O2 -5% CO2 by 95% N2 -5% CO2 for a similar period of time evoked changes which were for each ion in the same direction as those with GABA. In SR both GABA and N2 caused increases in [K+]o and decreases in [Cl-]o and [Na+]. The reduction of extracellular space, estimated from levels of [TMA+]o during exposures to GABA and N2, was 5-6% and insufficient to cause the observed changes in ion concentration. Ion changes induced by GABA and N2 were reversibly attenuated by the GABA(A) receptor antagonist bicuculline methiodide (BMI, 100 microM). GABA-evoked changes in [K+]o in SP and SR and [Cl-]o in SP were depressed by > or =90%, and of [Cl-]o in SR by 50%; N2-evoked changes in [K+]o in SP and SR were decreased by 70% and those of [Cl-]o by 50%. BMI blocked delta [Na+]o with both GABA and N2 by 20-30%. It is concluded that during early anoxia: (i) accumulation of GABA and activation of GABA(A) receptors may contribute to the ion changes and play a significant role, and (ii) responses in the dendritic (SR) regions are greater than and (or) differ from those in the somal (SP) layers. A large component of the [K+]o increase may involve a GABA-evoked Ca2+-activated gk, secondary to [Ca2+]i increase. A major part of [Cl-]o changes may arise from GABA-induced g(Cl) and glial efflux, with strong stimulation of active outward transport and anion exchange at SP, and inward Na+/K+/2Cl- co-transport at SR. Na+ influx is attributable mainly to Na+-dependent transmitter uptake, with only a small amount related to GABA(A) receptor activation. Although the release and (or) accumulation of GABA during anoxia might be viewed as potentially protectant, the ultimate role may more likely be an important contribution to toxicity and delayed neuronal death.     

Effect of GABA(B) receptor agonist on distension-sensitive pelvic nerve afferent fibers innervating rat colon

Spinal afferents innervating the gastrointestinal tract are the major pathways for visceral nociception. Many centrally acting analgesic drugs attenuate responses of visceral primary afferent fibers by acting at the peripheral site. Gamma-amino butyric acid (GABA), a major inhibitory neurotransmitter, acts via metobotropic GABA(B) and ionotropic GABA(A)/GABA(C) receptors. The aim of this study was to test the peripheral effect of selective GABA(B) receptor agonist baclofen on responses of the pelvic nerve afferent fibers innervating the colon of the rat. Distension-sensitive pelvic nerve afferent fibers were recorded from the S(1) sacral dorsal root in anesthetized rats. The effect of baclofen (1-300 micromol/kg) was tested on responses of these fibers to colorectal distension (CRD; 60 mmHg, 30 s). A total of 21 pelvic nerve afferent fibers was recorded. Mechanosensitive properties of four fibers were also recorded before and after bilateral transections of T(12)-S(3) ventral roots (VR). Effect of baclofen was tested on 15 fibers (7 in intact rats, 4 in rats with transected VR, and 4 in rats pretreated with CGP 54626). In nine fibers (5/7 in intact and 4/4 in VR transected rats), baclofen produced dose-dependent inhibition of response to CRD. Pretreatment with selective GABA(B) receptor antagonist CGP 54626 (1 micromol/kg) reversed the inhibitory effect of baclofen. Results suggest a peripheral role of GABA(B) receptors in the inhibition of mechanotransduction property of distension-sensitive pelvic nerve afferent fibers.     

Intracellular recording of the frog dorsal root single fibres' responses mediated by the GABAA and 5-HT-receptors

The effects of GABA, bicuculline and 5-HT on primary afferents in the isolated spinal cord of the frog Rana ridibunda were studied. Bath application of GABA (1 mM) reduced the primary afferent depolarisation (PAD) in IX segment of the spinal cord evoked by X dorsal root stimulation (57 +/- 8% of initial level, n = 5, p < 0.05). The action potentials (AP) recorded in dorsal root afferents was also suppressed under the GABA action (74 +/- 9%, p < 0.05). Bath application of bicuculline (50 microM) reduced the PAD (21 +/- 7%), n = 6, p < 0.05), meanwhile the AP in dorsal root afferents was resistant against the bicuculline action. Bath application of 5-HT (25 microM) depressed the PAD (34 +/- 7%, n = 7, p < 0.05) and the amplitude of the AP recorded from the single afferent fibre in dorsal column (76 +/- 6%, n = 7, p < 0.05). In contrast to GABA, 5-HT more effectively suppressed the late phase of the PAD evoked by X dorsal root stimulation and caused (76 +/- 6%, n = 7, p < 0.05) an alteration of the AP shape. All effects induced by these drugs were reversible. The mechanisms of GABA and 5-HT modulation of spinal cord afferent income are discussed.     

Barium spikes are generated in the spines of the sea urchin Diadema antillarum

In the presence of calcium, Ba2+ ions, at concentrations as low as 1-2 mM, block the action potentials (a.p.'s) elicited by the electrical stimulation of the primary spines of Diadema antillarum. Lower concentrations of barium (0.1 mM) potentiate the a.p.'s recorded from spines equilibrated with Ca-Free artificial sea water (ASW). Exposure of the spines to a saturated solution of EDTA in Ca-free, unbuffered ASW reversibly blocks their electrical activity. Spines blocked by EDTA continue to generate a.p.'s following their equilibration with Ca-free ASW containing 1 mM of Ba2+ ions. The time course of the a.p.'s recorded from spines equilibrated with normal ASW is only slightly affected by the combined action of 15 mM of tetraethylammonium (TEA) and 5 mM of 4-aminopyridine (4-AP). In contrast, the duration of the a.p.'s recorded from spines blocked by EDTA and placed in 1 mM barium is markedly increased by the combined actions of TEA and 4-AP at the above concentrations. We conclude that the Ca-channels of the neurites present in Diadema spines are not, at least qualitatively, an exception with regard to their permeation by Ba2+. The blocking action of low concentrations of these ions, particularly in the presence of calcium, may be explained by the extremely high surface to volume ratio prevailing in neurites with a diameter of only less than 0.1-0.3 micron.