Ionic mechanisms of the effect of phenibut and gaba not associated with a change in the function of the chloride channels

In experiments on isolated spinal cord of young rats 7–14 days old under conditions of takeoff of the electrical activity of the spinal roots with a sugar bridge, it was established that the GABA-mimetic phenibut induces direct depolarization of the motoneurons. In the same concentration range (10^–5-10^–4 M), GABA has a dual effect. The depolarizing component of the action of GABA in part of the experiments and the depolarizing effect of phenibut in all the experiments are preserved in the presence of picrotoxin (10^–5 M) and under conditions of superfusion of the brain with a solution with a reduced chloride concentration. This depolarizing effect of phenibut, not associated with the activation of GABA_A receptors and chloride channels coupled with them, is unchanged in a medium with Na⁺ deficiency, is enhanced during depolarization of the motoneurons due to an increased concentration of K⁺ (10 mM) and in the presence of imidazole, but is entirely eliminated in a medium with Ca²⁺ deficiency, containing 2 mM Mn²⁺, or in the presence of theophylline (10^–4 M). It is suggested that phenibut, and to some degree, GABA lower the intracellular concentration of cAMP by means of activation of the GABA_B receptors, which leads to blocking of the functional activity of the potential-dependent calcium channels and a decrease in the calcium-activated outflowing potassium currents. The ability to weaken the inflowing calcium currents may also be the basis of the presynaptic inhibiting effect of GABA and GABA-mimetics (phenibut, baclofen, etc.) on the pulsed release of mediators by the axon terminals of catecholaminergic, glutamatergic, and GABA-ergic neurons.A. M. Gor'kii Donetsk Medical Institute. Translated from Neirofiziologiya, Vol. 17, No. 4, pp. 481–489, July–August, 1985.     

Effect of serotonin on isolated cells with the various functionality from the lamprey spinal cord

The differential actions of 5-hydroxytryptamine (5-HT) (100 microM) were investigated on isolated motoneurons, interneurons, and primary sensory neurons from the lamprey spinal cord using patch-clamp techniques. Application of 5-HT did not evoke membrane currents in any of the spinal neurons tested (n = 62). However, in most motoneurons and interneurons (15 of 18), 5-HT produced a small depolarization (2-6 mV), which was not accompanied by a change in input resistance. In the remaining motoneurons and interneurons (3 of 18), 5-HT induced a large depolarization (up to 10-20 mV) and a decrease in input resistance of 20-60%. In most sensory neurons (dorsal sensory cells, DSCs), 5-HT evoked a short-lasting, low-amplitude depolarization, followed by a long-lasting hyperpolarization of 2-7 mV. The DSCs showed no significant change in input resistance to 5-HT application (n = 8). Spike afterpolarization were also differentially modulated by 5-HT. In motoneurons and interneurons, 5-HT decreased the amplitude of the afterhyperpolarization following the action potential while increasing the amplitude of the after depolarization. In the DSCs, no significant effect of 5-HT on spike afterpolarization was observed. 5-HT differentially modulated the current induced by application of N-methyl-D-aspartate (NMDA). In motoneurons and interneurons, 5-HT enhanced NMDA-evoked current, while in DSCs, 5-HT decreased this current. These results demonstrate that 5-HT differentially modulates the activity of functionally different groups of spinal neurons. In motoneurons and interneurons, 5-HT enhances excitation by inducing depolarization and decreasing the afterhyperpolatization, while NMDA currents are enhanced. These effects facilitate the appearance of rhythmic discharges in these cells in the presence of NMDA. In primary dorsal sensory cells, 5-HT enhances inhibition by hyperpolarizing the cells and depressing NMDA currents. These differential effects are presumably mediated by different types of 5-HT receptors on these classes of spinal neurons.     

Effects of 5-hydroxytryptamine on cat spinal motoneurons

The effects of 5-hydroxytryptamine (5-HT) on spinal motoneurons were examined in pentobarbital-anaesthetized cats and in nonanaesthetized decerebrate cats by intracellular recording and extracellular iontophoresis of 5-HT. 5-HT first induced a depolarization and then a long-lasting hyperpolarization (up to 60 min) with unchanged input resistance. The slow hyperpolarization was prevented by the 5-HT antagonists ketanserin (5-HT2), methysergide, and spiperone (5-HT1,2) and mimicked by the agonist (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (5-HT2). The post-spike after hyperpolarization was enhanced after application of 5-HT. A depolarization was induce by the 5-HT agonists (+/-)-8-hydroxy-(2)-(di-n-propylamino)tetralin (5-HT1A) and 1-(2-methoxyphenyl)piperazine (5-HT1). Possible mechanisms for the 5-HT-induced hyperpolarization and its intracellular medication are discussed. The present data suggest multiple effects of 5-HT on cat spinal motoneurons.     

GABA_A和GABA_B受体介导的蟾蜍背根神经节神经元胞体膜反应

实验在蟾蜍离体背根神经节(DRG)标本进行细胞内记录。浴槽滴加10~(-4)-10~(-3)mol/LGABA引起膜电位改变如下:(1)去极化(79/100);(2)双相反应;先为去极化,继后为超极化(10/100);(3)无反应(11/100)。以上去极化反应均可为荷包牡丹碱所阻断。GABA-去极化时膜电导增加,逆转电位值为-15——25mV。低Cl~-和高Cl~-任氏液分别使GABA-去极化反应增大和减小。10~(-4)mol/Lbaclofen不引起膜电位改变。在GABA-去极化期间,观察到大部分细胞的动作电位时程(ApD)缩短。ApD的此种变化可为baclofen所模拟,但不为荷包牡丹碱所阻断。结果提示:蟾蜍DRG神经元胞体膜有GABA_A和GABA_B受体共存,前者介导膜电位的改变,后者介导ApD的缩短。本文并联系到初级传入终末的突触前抑制的产生机制进行了讨论。     

The effects of acetylcholine, carbamylcholine and gamma-aminobutyric acid on uropod motoneurons in the crayfish Procambarus clarkii and Cambaroides japonicus

The effects of acetylcholine (ACh), carbamylcholine and gamma-aminobutyric acid (GABA) on the spike activity of uropod motoneurons were investigated electrophysiologically in the crayfish Procambarus clarkii Girard and Cambaroides japonicus de Haan. High concentrations of ACh were required to bring about an increase in the spike discharge of uropod motoneurons while carbamylcholine, which is not destroyed by cholinesterase, caused a marked increase in the motoneuron spike discharge even in low concentrations. Application of GABA in concentrations of 10(-5)-10(-2) M caused the decrease in the spike discharge of uropod motoneurons. Under the condition that the synaptic transmission onto uropod motoneurons was blocked by perfusing EGTA containing Ca2+-free saline with high-Mg2+, ACh increased the spike discharge of uropod motoneurons whereas GABA decreased it. The results suggested that ACh and GABA function as excitatory and inhibitory transmitters, respectively, in the crayfish central nervous system.     

Effect of gamma-aminobutyric acid and phenibut on the central links of the vascular reflexes with chemo- and mechanoreceptors of the aorto-carotid zone]

Experiments were conducted on decerebrated cats. A depressive effect of gamma-aminobutyruc acid (GABA)--100--200 mg/kg and its phenyl derivative phenibut--20 mg/kg--on depressive reactions of the systemic arterial pressure and on the inhibition of spontaneous bioelectrical activity in the renal nerve occurring in stimulation of the mechanoreceptors of the carotid sinus and of the sinus and depressor nerve afferents (having a mechanoreceptor modality) was demonstrated. Pressor reactions of the systemic arterial pressure and evoked bioelectrical activity were enhanced in the renal nerve in stimulation of chemoreceptors of the carotid sinus following administration of the same GABA and phenibut doses. The data obtained are interpreted from the aspect of a deprimating action of GABA and phenibut in the area of the paramedian reticular nuclei of the medulla oblongata.     

Spinal ganglion neurons of rats--a model for the study of the central serotonin receptors

Application of 5-hydroxytryptamine (5-HT) (3 x 10(-5) M) on the rat lumbar dorsal ganglia (RDG) induced membrane depolarization with increased input resistance in 30% of neurons, hyperpolarization with decreased input resistance in 30% of neurons and mixed responses in 40% of neurons. Methysergide and amitriptyline (10(-6) M) blocked depolarizing but not hyperpolarizing effects of 5-HT. Propranolol (3 x 10(-6) M) was inactive in respect to both 5-HT responses. 5-HT depolarizing responses of RDG neurons were mediated by 5-HT2 receptors activation and decreased membrane potassium conductivity; 5-HT hyperpolarizing responses were mediated by 5-HT1A receptor activation and increased potassium conductivity. RDG neurons seem to be an interesting model for the investigation of central 5-HT receptor mechanism.     

Neutrophil hyperpolarization in response to a chemotactic peptide

The chemotactic peptide formylmethionyl-leucyl-phenylalanine (fMLP), at concentrations below 10(-9) M, elicits a sustained increase in the human neutrophil's membrane potential within 10 s of its addition. This hyperpolarization, detected with the fluorescent cationic potentiometric probes, 3,3'-dipentyloxacarbocyanine (diO-C5-(3)), and 1,1'-dipropyl-3,3,3',3'-tetramethylindocarbocyanine iodide (diI-C3-(3)), and with the anionic probe bis-(1,3-diethylthiobarbituric)trimethine oxonol (bis-oxonol), is immediately followed by a large depolarization when [fMLP] greater than 10(-9) M. By extracellular substitution of sodium ions with potassium ions or choline or by pretreatment of the cells with ionophores, we report here that the hyperpolarization is primarily dependent on an intact potassium ion gradient and is accompanied by a concurrent acidification of the cytoplasm (approximately 0.05 pH unit) Although the latter occurs simultaneously with a large, transient increase in cytosolic Ca2+ at [fMLP] greater than 10(-10) M, it occurs without a detectable increase in cytosolic Ca2+ at [fMLP] less than 10(-10) M. The hyperpolarization is neither affected nor initiated by the chemotactic peptide antagonist tert-butyloxycarbonyl-methionyl-leucyl-phenylalanine, whereas the depolarization is completely inhibited. Neutrophils isolated from patients with X-linked chronic granulomatous disease exhibit normal hyperpolarizations and cytosolic Ca2+ increases in response to chemotactic peptides but exhibit no depolarization or oxidative burst. The hyperpolarization appears earlier in the ontogeny of differentiating myeloid precursor cells than either the rise in cytosolic Ca2+ or the depolarization response. Together, these findings indicate that an increase in transmembrane potential is one of the earliest events in the neutrophil response to chemotactic peptides, coinciding temporally with increases in cytoplasmic Ca2+ and H+ concentrations but preceding detectable oxidative burst activity.     

Veratridine-induced oscillations in membrane potential of cultured rat skeletal muscle: Role of the Na-K pump

1. The acute effects of veratridine on membrane potential (Em) and Na-K pump activity in cultured skeletal muscle were examined. 2. At a concentration of 10(-4) M, veratridine caused depolarization of Em and a decrease in Na-K pump activity. At concentrations of 10(-5) and 10(-6) M, veratridine caused oscillations of Em and an increase in Na-K pump activity compared to untreated, control cells. The oscillations consisted of depolarization to about -40 mV followed by hyperpolarization to about -90 mV; the level of hyperpolarization was higher at 37 than at 23 degrees C. 3. Veratridine-induced oscillations could be prevented by pretreatment with tetrodotoxin (10(-6) M) and blocked or prevented by ouabain, which depolarizes Em of cultured myotubes. In contrast, depolarization of Em to -60 mV by excess K+ did not alter the amplitude or frequency of the oscillations. 4. The results demonstrate that veratridine-induced increase in Na influx both depolarizes cultured myotubes and increases the activity of the Na-K pump, which repolarizes Em to levels higher than control. This sequence accounts for veratridine-induced oscillations in Em. High concentrations of veratridine cause only depolarization of Em and inhibition of Na-K pump activity.     

Competitive antagonism of insect GABA receptors by iminopyridazine derivatives of GABA

A series of 4-(6-imino-3-aryl/heteroarylpyridazin-1-yl)butanoic acids were synthesized and examined for antagonism of GABA receptors from three insect species. When tested against small brown planthopper GABA receptors, the 3,4-methylenedioxyphenyl and the 2-naphthyl analogues showed complete inhibition of GABA-induced fluorescence changes at 100 μM in assays using a membrane potential probe. Against common cutworm GABA receptors, these analogues displayed approximately 86% and complete inhibition of GABA-induced fluorescence changes at 100 μM, respectively. The 4-biphenyl and 4-phenoxyphenyl analogues showed moderate inhibition at 10 μM in these receptors, although the inhibition at 100 μM was not complete. Against American cockroach GABA receptors, the 4-biphenyl analogue exhibited the greatest inhibition (approximately 92%) of GABA-induced currents, when tested at 500 μM using a patch-clamp technique. The second most active analogue was the 2-naphthyl analogue with approximately 85% inhibition. The 3-thienyl analogue demonstrated competitive inhibition of cockroach GABA receptors. Homology modeling and ligand docking studies predicted that hydrophobic 3-substituents could interact with an accessory binding site at the orthosteric binding site.     

Effect of inhibitory amino acid antagonists on postsynaptic potentials of motoneurons of the frog Rana ridibunda

On preparations of the isolated spinal cord of the frog Rana ridibunda at intracellular recording from lumbar motoneurons, it is shown that response to the 10 mM GABA application decreased selectively by 40.7 ± 23.7% (n = 6) as a result of the spinal cord treatment with bicuculline (100–150 μM), while response to the Gly application decreased selectively by 50.7 ± 17.8% (n = 10) after the spinal cord treatment with strychnine (5–10 μM). Both strychnine and bicuculline produced potentiation of EPSP by amplitude and duration as well as paroxysmal depolarizational shifts (PDS). Strychnine produced more effectively the potentiation, while bicuculline—PDS. The inhibitory Gly effect decreased significantly the DR and RF EPSP (a decrease of amplitude and duration) as a result of the spinal cord treatment with strychnine (5–10 μM), but not with bicuculline. The inhibitory GABA effect on the DR and RF EPSP decreased as a result of the spinal cord treatment with bicuculline only in a half of the studied motoneurons and to the lesser degree than the inhibitory Gly effect on the same EPSP at the strychnine treatment. Based of the obtained data, it is suggested that the inhibitory effects on the excitatory inputs of the motoneuron in the frog are expressed weaker than in mammals and are related predominantly to the GABA and Gly effects on receptors of interneurons. It is suggested that GABA specifically acts mostly on GABA_A receptors, whereas Gly—on Gly receptors, although there is some part of cross-inhibition. Original Russian Text ? G. G. Kurchavyi, N. I. Kalinina, and N. P. Vesselkin, 2006, published in Zhurnal Evolyutsionnoi Biokhimii i Fiziologii, 2006, Vol. 42, No. 5, pp. 463–471. The present work is a continuation of the work [1].     

Thyrotropin-releasing hormone mimics descending slow synaptic potentials in rat spinal motoneurons

Thyrotropin-releasing hormone (TRH) produced a depolarization in lumbar motoneurons of neonatal rats. The depolarization by TRH persisted after extracellular Ca2+ was replaced by Mg2+ or Mn2+, indicating its direct action upon motoneurons. Stimulation of the ventral descending tract at the lower thoracic segment evoked slow excitatory postsynaptic potentials (e.p.s.ps) lasting 20-30 s in every motoneuron. Both the TRH-induced depolarization and descending slow e.p.s.p. were accompanied by a decrease in input conductance of motoneurons. When the membrane potential of the motoneuron was shifted, both the TRH-induced depolarization and slow e.p.s.p. became larger in amplitude during depolarization and smaller during hyperpolarization. However, they could not be reversed in polarity by hyperpolarization. During the depolarization of motoneuron produced by TRH application, the slow e.p.s.p. was markedly reduced in amplitude, suggesting the involvement of identical ionic mechanisms in the two responses. After incubation of the isolated spinal cord with antisera to TRH, the depolarizing response produced by TRH as well as the descending slow e.p.s.p. was greatly diminished. In contrast, monosynaptic reflexes evoked by dorsal root stimulation remained unchanged under this condition. These results suggest that TRH serves as a neurotransmitter mediating the descending slow e.p.s.p. in motoneurons.     

Functional assay for GABA receptor subtypes of a cockroach giant interneuron.

gamma-Aminobutyric acid (GABA) receptors were examined in the cockroach central nervous system (CNS) using the single fiber-oil gap method applied to an identified giant interneuron. Short-lasting pressure application of 10 mM GABA developed a multiphasic response composed of a fast hyperpolarization followed by a transient depolarizing component and a stable hyperpolarization. This triphasic characteristic shape of the response was modified according to the dose of GABA injected or bath-applied and to the precise localization of the injection within the dendritic area. The transient depolarizing phase showed a negative reversal potential of -70 mV. Both hyperpolarizing phases reversed at a more negative level ranging to -80 mV. A positive shift of these values was caused by a decrease in external chloride concentration. Bath-application of 0.1 mM picrotoxin (Ptx) decreased the depolarizing phase which was progressively replaced by a stable hyperpolarization. The transient depolarizing component desensitized quickly and was the most sensitive phase to Ptx action. The Ptx-resistant response reversed at a mean value of -100 mV close to the equilibrium potential for potassium ions (EK+), suggesting that it was generated by a K(+)-channel coupled receptor. Although baclofen was unable to mimic the Ptx-resistant GABA response, the compound CGA 147823, known to bind with a high specificity to vertebrate GABAB receptors, has been successfully used to reproduce the Ptx-resistant GABA response. It is suggested that, in addition to GABA receptors linked to chloride channels, the insect CNS possesses GABA receptors sharing ionic characteristics of GABAB receptors especially those located in the vertebrate CNS, although they are insensitive to baclofen.     

Effects of orexins/hypocretins on neuronal activity in the paraventricular nucleus of the thalamus in rats in vitro

Orexin-A (ORX-A) and orexin-B (ORX-B), also called hypocretin-1 and hypocretin-2, respectively, act upon orexin 1 (OX1R) and orexin 2 (OX2R) receptors, and are involved in the regulation of sleep-wakefulness and energy homeostasis. Orexin neurons in the lateral hypothalamic perifornical region project heavily to the paraventricular nucleus of the thalamus (PVT), which is deeply involved in the control of motivated behaviors. In the present study, electrophysiological and cytosolic Ca2+ concentration ([Ca2+]i) imaging studies on the effects of ORX-A and ORX-B on neurons in the PVT were carried out in rat brain slice preparations. ORX-A and/or ORX-B were applied extracellularly in the perfusate. Extracellular recordings showed that about 80% of the PVT neurons were excited dose-dependently by both ORX-A and ORX-B at concentrations of 10(-8) to 10(-6)M, and the increase in firing rate was about three times larger for ORX-B than for ORX-A at 10(-7)M. When both ORX-A and ORX-B were applied simultaneously at 10(-7)M, the increase in firing rate was almost equal to that of ORX-B at 10(-7)M, suggesting that the PVT neurons do not show a high affinity to ORX-A which is expected if they have OX1R receptors. The excitatory effect of ORX-B was seen in low Ca2+ and high Mg2+ ACSF as well as in normal ACSF, and the increase in firing rate was greater in low Ca2+ and high Mg2+ ACSF than in normal ACSF. [Ca2+]i imaging studies demonstrated that [Ca2+]i was increased in about 50% of the PVT neurons by both 10(-7)M ORX-A and ORX-B with a stronger effect for ORX-B, and the increase in [Ca2+]i induced by ORX-B was abolished in Ca2+-free ACSF, suggesting that ORX-B does not release Ca2+ from intracellular Ca2+ stores. Subsequent whole cell patch clamp recordings revealed that an after hyperpolarization seen following each action potential in normal ACSF disappeared in Ca2+-free ACSF, and the mean magnitude of the depolarization induced by ORX-B was same in normal, Ca2+-free and TTX-containing Ca2+-free ACSFs. Furthermore, ORX-B-induced depolarization was reversed to hyperpolarization when membrane potential was lowered to about -97 mV, and an increase of extracellular K+ concentration from 4.25 to 13.25 mM abolished the ORX-B-induced depolarization, indicating that the ORX-B-induced depolarization is associated with an increase in the membrane resistance resulting from a closure of K+ channels. These results suggest that orexins depolarize and excite post-synaptically PVT neurons via OX2R receptors, and that orexin-activated PVT neurons play a role in the integration of sleep-wakefulness and energy homeostasis, and in the control of motivated behaviors.     

Characteristics of GABA receptors on the ocellar L-neurons of American cockroach Periplaneta americana

The ocellar L-neurons of cockroach Periplaneta americana were used in the present study as model systems to investigate the pharmacological properties of the GABA receptors. To do so, a glass microelectrode was impaled into the axon of the L-neurons to record the membrane potential intracellularly and to monitor membrane response to GABA treatment and cercal stimulation by air puff. The traditional GABA and their receptor agonists were introduced through perfusion and/or iontophoresis to monitor their effects on the L-neurons. The GABA receptor antagonists were administered by perfusion to examine if the response of the L-neurons to GABA and/or cercal stimulation was changed. The results revealed that administration of GABA, muscimol and imidazole acetic acid, two GABAA agonists, produced depolarization on the L-neurons. However, treatment of 3-APS and guanidine acetic acid, another two GABAA agonists, evoked hyperpolarization on the L-neurons. Among those tested antagonists, only picrotoxin, GABAA antagonist, antagonize the depolarization induced by GABA and/or cercal stimulation. More interestingly, administration of strychnine, glycine receptor antagonist, largely attenuated the depolarization response of the L-neurons to cercal stimulation. This attenuation caused by strychnine was even stronger than that initiated by varied GABA antagonists. In addition, phaclofen, a GABAB receptor antagonist, showed no antagonistic effect. These results strongly suggest that the characteristics of GABA receptors of the ocellar L-neurons may differ from those in vertebrates. It may be more likely to be a novel GABA receptor.     

Acetylcholine receptors in monkey and rabbit oocytes

Membrane potential responses to acetylcholine (ACh, 10(-7)-10(-3 M) were investigated in monkey and rabbit ovarian oocytes. In monkey oocytes ACh most commonly elicited a short-latency hyperpolarization concomitant with a decreased membrane input resistance (Rin). Under voltage-clamp short-latency ACh currents had an equilibrium potential of approximately -40 mV. In rabbit oocytes responses to ACh consisted of an increase in Rin or of a depolarization with an equilibrium potential of approximately -15 mV. Curare, hexamethonium, and atropine (10(-5)-10(-3) M) did not block these ACh responses. Thus, the oocyte membrane in the rabbit contains ACh receptors that cannot be classified as either muscarinic or nicotinic.     

Nitric oxide and endothelin in oxygen-dependent regulation of vascular tone of human umbilical vein

We investigated the possible contribution of nitric oxide (NO) and endothelin (ET) to oxygen-dependent regulation of human umbilical vein vascular tone by simultaneous registration of intracellular membrane potential and isometric tension of vessel strips with and without NO synthase inhibition [10-4 M N omega-nitro-L-arginine methyl ester (L-NAME)], ETA receptor blockade (10(-5) M BQ-123), or ETB receptor blockade (10(-7) M BQ-788) at Po2 values in the bath solution between 5 and 104 mmHg. Increasing PO2 above the physiological intrauterine range resulted in depolarization and an increase of isometric tension, whereas lowering PO2 resulted in hyperpolarization and a decrease in isometric tension. Removal of the endothelium reversed these effects. At PO2 values below 39 mmHg, intact preparations treated with either L-NAME, BQ-788, or BQ-123 were more depolarized than controls. In the case of treatment with L-NAME or BQ-123, this was accompanied by an increase in isometric tension. We conclude that it is NO that mediates the hypoxic hyperpolarization and vasodilatation of the human umbilical vein and that ET, via activation of ETB1 receptors on endothelial cells, contributes to this effect.     

GABA receptors on the cell-body membrane of an identified insect motor neuron

The pharmacology of a gamma-aminobutyric acid (GABA) receptor on the cell body of an identified motor neuron of the cockroach (Periplaneta americana) was investigated by current-clamp and voltage-clamp methods. Iontophoretic application of GABA increased membrane conductance to chloride ions, and prolonged application resulted in desensitization. Hill coefficients, determined from dose-response data, indicated that binding of at least two GABA molecules was required to activate the chloride channel. Differences between vertebrate GABAA receptors and insect neuronal GABA receptors were detected. For the GABA receptor of motor neuron Df, the following rank order of potency was observed: isoguvacine greater than muscimol greater than or equal to GABA greater than 3-aminopropanesulphonic acid. The GABAB receptor agonist baclofen was inactive. Of the potent vertebrate GABA receptor antagonists (bicuculline, pitrazepin, RU5135 and picrotoxin), only picrotoxin (10(-7) M) produced a potent, reversible block of the response to GABA of motor neuron Df. Both picrotoxinin and picrotin also blocked GABA-induced currents. Bicuculline hydrochloride (10(-4) M) and bicuculline methiodide (10(-4) M) were both ineffective when applied at resting membrane potential (-65 mV), although at hyperpolarized levels partial block of GABA-induced current was sometimes observed. Pitrazepin (10(-4) M) caused a partial, voltage-independent block of GABA-induced current. The steroid derivative RU5135 was inactive at 10(-5) M. In contrast to the potent competitive blockade of vertebrate GABAA receptors by bicuculline, pitrazepin and RU5135, none of the weak antagonism caused by these drugs on the insect GABA receptor was competitive. Flunitrazepam (10(-6) M) potentiated GABA responses, providing evidence for a benzodiazepine site on an insect GABA-receptor-chloride-channel complex.     

Activation of picrotoxin-resistant GABA receptors by GABA and related compounds induces modulation of cockroach dorsal paired median (DPM) neuron firing

Activation of gamma-aminobutyric acid (GABA) receptors in insect dorsal paired median (DPM) neurons induced two types of response which appeared to be mediated by two different GABA receptor subtypes. When activated by bath application of GABA, one receptor subtype, insensitive to picrotoxin (PTX), mediated a drastic reduction in the firing frequency, leading to a blockade of the spontaneous electrical activity. These effects were accompanied by decreases in the amplitude and duration of the plateau action potential (AP) and the spike after-hyperpolarization (AHP). In most cases, a slight depolarization of the resting membrane potential occurred. Bath application of the vertebrate GABA(B) receptor agonists 3-aminopropyl(methyl)phosphinic acid (SKF 97541) and 3-aminopropylphosphinic acid (CGA 147823/CGP 27492) induced similar responses. Another GABA receptor subtype, less sensitive to GABA, mediated a chloride dependent hyperpolarization that was suppressed by bath application of PTX. The approximate locations of these two GABA receptor subtypes were determined by local pressure microapplications of GABA and vertebrate GABAergic agonists. The PTX-sensitive receptors were located predominantly on the surface of the ganglion where the apical pole of the soma is situated, while the PTX-resistant receptors appeared to be located deeper within the ganglion.These results reveal the existence of two GABA receptor subtypes on the DPM neurons and provide evidence for a functional role for PTX-resistant GABA receptors in the regulation of spontaneous firing.     

Inhibitory and excitatory mechanisms of neurotensin action in canine intestinal circular muscle in vitro.

The effect of neurotensin on canine ileal circular muscle devoid of myenteric plexus was investigated using single and double sucrose gap techniques. Similar results were obtained with microelectrode techniques. Neurotensin caused a temperature-sensitive and dose-dependent biphasic response, an initial hyperpolarization associated with inhibition of contractile activity, followed by an excitatory phase, usually consisting of spike discharge and tonic and phasic contractions, for which depolarization was not required. Neither response was affected by tetrodotoxin, phentolamine, propranolol, or atropine. The hyperpolarization was associated with decreased membrane resistance, blocked by 10(-7) M apamin, and converted to tonic depolarization by apamin (10(-6) M). Tachyphylaxis to neurotensin occurred when the stimulation interval was less than 20 min. After Ca2+ depletion, depolarization was observed instead of the hyperpolarization; this depolarization was not affected by nitrendipine and was gradually abolished with repetitive stimulation at 20-min intervals. When Ca2+ was present, nifedipine did not alter the hyperpolarizing phase of the response but inhibited spiking and blocked all contractions. The excitatory phase of the response was enhanced by Bay K-8644. Neuromedin N elicited a response identical with that of neurotensin. The responses of the two peptides were completely cross tachyphylactic. Inhibitory junction potentials were not affected by neurotensin tachyphylaxis. It is concluded that neurotensin and neuromedin N activate apamin-sensitive, calcium-dependent potassium channels in circular muscle, causing membrane hyperpolarization and inhibition of muscle contraction. Release of intracellular calcium is involved in the activation of these potassium channels.(ABSTRACT TRUNCATED AT 250 WORDS)