Increase In Membrane Resistance Research Articles

Triggered Activity Induced by K+ -Free, Na+-Deficient Solution in Guinea Pig Ventricular Muscle

Triggered activity induced by delayed afterdepolarizations has been suggested as a cellular mechanism for some arrhythmias. The development of triggered activity should be favored by conditions that increase myoplasmic Ca2+ and increase membrane resistance. A solution which could create these conditions was devised. After perfusion with normal Tyrode's solution, 24 trabeculae were exposed to the experimental solution. All preparations developed triggered activity after exposure to the experimental solution, 83% developed delayed afterdepolarizations before the onset of triggered activity, and triggered activity stopped in all trabeculae after reperfusion with normal Tyrode's solution. The interaction of ouabain, lidocaine, and three Ca2+ channel blockers with triggered activity induced by the experimental solution is described. Verapamil inhibited triggered activity but not underlying voltage oscillations, whereas nisoldipine and Mn2+ inhibited both triggered activity and voltage oscillations. Lidocaine did not inhibit afterdepolarizations or triggered activity. Exposure to ouabain for 10 min caused delayed afterdepolarizations but not triggered activity. Our results show that the experimental solution induced triggered activity, which was highly reliable and readily reversible. The high reproducibility of this activity enables the study of interactions of pharmacological agents with this triggered activity. This may contribute to the further understanding of the mechanism underlying some arrhythmias.

Phosphatidylserine restores spatial memory and morphofunctional cholinergic markers in basal forebrain nuclei of aged rats : ∗M.G. Nunzi, F. Milan, P. Polato, A. Zanotti, D. Guidolin and G. Toffano. Fidia Research Laboratories, 35031 Abano Terme (PD), Italy

The effects of disulfide bond and sulfhydryl group reagents on synaptic transmission were studied in the bullfrog sympathetic ganglion. Ten millimoles of dithiothreitol (DTT). a disulfide bond reducing agent, decreased the amplitude of the transmitted postganglionic compound action potential (CAP) to 41% of control. More than one hour wash-out of DTT with normal Ringer's solution did not reverse the block, but 1 mM 5.5'-dithiobis(2-nitrobenzoic acid) (DTNB), an oxidizing agent, rapidly restored the compound action potential amplitude. N-Ethylmalemide (NEM), an alkylating agent, added before DTNB, prevented the restoration of transmission by DTNB. Block of transmission induced by dithiothreitol was also reversed rapidly by 50 μm 3,4-diaminopyridine (3,4-DAP), to 87% of control compound action potential amplitude, and subsequent addition of DTNB restored transmission completely and this was accompanied by the stimulus-bound repetitive firing (SBR) which diaminopyridine produced in otherwise untreated ganglia.One to 3 mM dithiothreitol decreased the amplitude of excitatory post-synaptic potentials (EPSP), and greater concentrations slightly depolarized the cell membrane and decreased membrane resistance. Washout with Ringer's solution reversed the depolarization, but not the block of excitatory postsynaptic potentials. The amplitude of excitatory post-synaptic potentials was restored to threshold by 1 mM DTNB, which also slightly increased membrane resistance and the resting potential.It is concluded that the actions of dithiothreitol and DTNB in the bullfrog sympathetic ganglion are generally similar to those previously observed in other junctional tissues. Interactions between these drugs and 3,4-diaminopyridine in the present study also raise the question whether dithiothreitol and DTNB have any presynaptic effects.

Membrane resistance increase induced in thalamic neurons by stimulation of brainstem cholinergic afferents

The synaptic response induced in thalamic relay cells by stimulation of peribrachial cholinergic afferents was studied in cats maintained under urethane anesthesia. Recordings were performed in the ventral lateral nucleus after destruction of the reticular thalamic complex and cortical lesion. Some animals were also pretreated with reserpine (1.2 mg/kg) 24 h before recording sessions. Repetitive stimulation (25–60 Hz for 200–600 ms) of the peribrachial area produced a slow depolarization and tonic firing in thalamic cells. The depolarization was associated with an increase in membrane resistance and its amplitude decreased during membrane hyperpolarization. The reactivity of thalamic relay cells to a corticothalamic volley was also increased during the slow depolarization. Similar results were obtained in reserpinized animals. On the basis of previous data obtained in vitro, it is concluded that the slow depolarization resulted from the activation of muscarinic receptors by brainstem cholinergic afferents.

Phase dependency of electrotonic spread of hyperpolarizing current pulses in the rabbit sinoatrial node

Electrotonic current spread in the SA node of the rabbit was measured by means of hyperpolarizing current pulses (1 to 10 μA, 60 ms), which were injected intracellularly through a K +-perfused suction electrode. The pulses were applied at the beginning, middle or end of the diastolic depolarization phase. The resulting membrane potential change of nodal fibers was measured with microelectrodes. Space constants were calculated by fitting single exponential curves to the data. The input resistance ( R in) of fibers at different sites in the SA node was measured by means of a double barrel microelectrode (current pulses 5.5 to 11 nA, 60 ms) to detect a change in the internal resistance during the diastolic depolarization phase. During diastole the average electrotonic potential increased by 30% ( P < 0.001), the increase of the space constant ranged from 9 to 183% ( P < 0.05). R in however, did not change during diastole. It is concluded that the electrotonic spread increased phase dependently, due to an increase of membrane resistance; the internal resistance was not phase dependent.

Resistance measurements as a simple diagnostic tool for ion‐selective electrode performance

AbstractRecently, potentiometric sensors have found numerous applications involving on‐line monitoring in flow systems [1–4]. With trends toward the use of multielectrode arrays and electrode miniaturization, simple diagnostic tests for the routine assessment of device performance will become increasingly important. In this work, we present the possibility of using dc resistance measurements for this purpose. Mini‐sodium‐selective electrodes with polyvinyl chloride (PVC) membranes incorporating the ionophore methyl p‐t‐butylcalixaryl acetate [4, 5] were constructed, and a gradual increase in resistance with lifetime was observed. A lifetime mimicking study was set up in which leaching of the active components from the membrane was simulated by a series of dilutions, and the effect on overall electrode performance was monitored at each stage. Complete electrode malfunction has been shown to occur abruptly when the active membrane component concentration falls below a critical level. Electrode failure is accompanied by a sharp increase in membrane resistance.

Membrane resistance increases when automaticity develops in explanted rat heart cells

We compared the passive electrical properties of isolated ventricular myocytes (resting potential -65 mV, fast action potentials, and no spontaneous activity) with those of 2- to 7-day-old cultured ventricle cells from neonatal rats (resting potential -50 mV, slow action potentials, and presence of spontaneous activity). In myocytes the specific membrane capacity was 0.99 microF/cm2, and the specific membrane resistance increased from 2.46 k omega.cm2 at -65 mV to 7.30 k omega.cm2 at -30 mV. In clusters, the current-voltage relationships measured under current-clamp conditions showed anomalous rectification and the input resistance decreased from 1.05 to 0.48 M omega when external K+ concentration was increased from 6 to 100 mM. Using the model of a finite disk we determined the specific membrane resistance (12.9 k omega.cm2), the effective membrane capacity (17.8 microF/cm2), and the lumped resistivity of the disk interior (1,964 omega.cm). We conclude that 1) the voltage dependence of the specific membrane resistance cannot completely explain the membrane resistance increase that accompanies the appearance of spontaneous activity; 2) a decrease of the inwardly rectifying conductance (gk1) is mainly responsible for the increase in the specific membrane resistance and depolarization; and 3) approximately 41% of the inward-rectifying channels are electrically silent when spontaneous activity develops in explanted ventricle cells.

Involvement of a cholinergic mechanism in the sustained depolarization and contraction of the lower oesophageal sphincter muscle cells in the cat

Membrane potentials were recorded in vitro with intracellular electrodes from the circular muscle cells of the cat lower oesophageal sphincter and oesophageal body. In addition, the tension of lower oesophageal sphincter and oesophageal body strips was recorded isotonically. Under the experimental conditions, no spontaneous electrical activity or variation in the tension of the strips occurred. The resting membrane potential of the circular muscle cells was significantly lower in the lower oesophageal sphincter (−51.0 ± 0.3 mV) than in the circular muscle cells of the oesophageal body (−57.1 ± 0.4 mV). These values were not affected by infusion of tetrodotoxin 3.1 × 10 −6M. In the presence of atropine (3.5 × 10 −7M), the resting membrane potential of the circular muscle cells of the lower oesophageal sphincter increased significantly (−57.6 ± 0.4 mV), whereas the resting membrane potential of the circular muscle cells of the oesophageal body was not significantly affected (−57.8 ± 0.6 mV). In the presence of atropine, no significant difference in the values of the resting membrane potential of the circular muscle cells was observed between the lower oesophageal sphincter and the oesophageal body. Hyoscine (2.9 × 10 −7M) significantly increased the resting membrane potential of the circular muscle cells of the lower oesophageal sphincter, whereas eserine (3.6 × 10 −6M) significantly decreased it. Atropine induced a significant decrease in the membrane resistance of the circular muscle cells of the lower oesophageal sphincter. Atropine decreased the resting tension of lower oesophageal sphincter strips whereas eserine increased it, but no such effects were recorded on oesophageal body strips. These results suggest that, in the cat, the circular muscle cells of the lower oesophageal sphincter, but not those of the oesophageal body, are continuously influenced by cholinergic nerve terminals, which cause a low resting membrane potential, an increase in membrane resistance and persistent contraction. These effects of cholinergic neurons are not action potential dependent. The results also show that the sustained depolarization of the circular muscle cells of the lower oesophageal sphincter elicits persistent contraction of these cells. This mechanism may play a key role in the persistent closure of this sphincter.

Electrical properties of canine subendocardial Purkinje fibers surviving in 1-day-old experimental myocardial infarction.

The passive electrical properties of subendocardial Purkinje fibers surviving in infarcted regions of canine ventricle 24 hours after coronary ligation were studied by using microelectrode techniques and cable theory. In normal hearts, cells within the subendocardial Purkinje fiber strands were found to be well coupled to each other but electrically isolated from neighboring myocardium. Voltage response to intracellular current injection was consistent with one-dimensional cable behavior and yielded estimates of passive electrical properties in general agreement with previous work on free-running Purkinje strands (membrane length constant, 1.2 +/- 0.1 mm; membrane time constant, 7.3 +/- 0.8 msec; input resistance, 67.4 +/- 7.4 K omega; membrane resistance, 8.2 +/- 0.7 K omega.cm; axial resistance, 0.52 +/- 0.06 M omega/cm; membrane capacitance, 960 +/- 102 nF/cm) (n = 21). On the day after coronary ligation, subendocardial Purkinje fiber action potentials were prolonged and slightly depolarized. Significant increases were measured in input resistance (+40.5%), membrane resistance (+43.9%), and axial resistance (+47.5%), whereas membrane capacitance was found to be significantly decreased (-24.3%) (n = 19). Conduction velocity, membrane length constant, membrane time constant, and the time constant and capacitance for the foot of the action potential remained unchanged. These results are consistent with electrical uncoupling between adjacent cells, which will increase internal resistivity, accompanied by changes in cellular phospholipid content, which can increase membrane resistance and alter membrane capacitance. Alternatively, the results can be explained by a simple model in which the apparent electrical structure is altered by changes in electrical coupling alone, with specific electrical properties remaining constant. Although the mechanisms underlying the observed changes remain uncertain, the present study indicates that myocardial infarction is associated with alterations in the passive electrical structure of surviving subendocardial Purkinje fibers, which, together with changes in action potential configuration, may provide a substrate for the generation of ventricular arrhythmias 24 hours after coronary ligation.

Excitation of hippocampal neurons by stimulation of glutamate Q p receptors

The electrophysiological effects of Qp receptors activation in brain neurons are investigated in rat hippocampal slice preparation. Quisqualate, in the presence of a ionotropic quisqualate receptor antagonist that does not block Qp receptor activation, produces a depolarization associated with an increased membrane resistance. All results indicate that the recently identified glutamate Qp receptor subtype may play an important role in the regulation of neuronal excitability by excitatory amino acid transmitters

Excitatory and inhibitory effects of epinephrine on neonatal rat sympathetic preganglionic neurons in vitro

Current and voltage recordings were made from antidromically identified sympathetic preganglionic neurons (SPNs) in transverse thoracolumbar spinal cord slices removed from neonatal rats. When applied by either pressure ejection or superfusion, epinephrine (Epi) caused a slow depolarization or an inward current in 62 SPNs (42%) and a slow hyperpolarization or an outward current in 21 SPNs (14%). The responses persisted in low calcium- or tetrodotoxin-containing media. The Epi-induced depolarization or inward current was associated with increased membrane resistance; it was reduced by membrane hyperpolarization and nullified at a membrane potential of about −100 mV; a clear reversal however was not observed at more negative potential levels. In a number of SPNs the Epi-induced depolarization was accompanied by small inhibitory postsynaptic potentials. The latter were eliminated by a low calcium solution and by the glycine antagonist strychnine, suggesting that they were caused by glycine or a glycine-like substance released from interneurons subsequent to activation by Epi. The Epi-induced hyperpolarization or outward current was associated with decreased membrane resistance, and nullified around −100 mV. The α-adrenergic antagonist, dihydroergotamine, andα 1-antagonist, prazosin, reversibly blocked the excitatory, whereas theα 2-antagonist, yohimbine, abolished the inhibit response, respectively. It is concluded that Epi acting onα 1-andα 2-adrenergic receptors depolarizes and hyperpolarizes the r SPNs by decreasing or increasing membrane conductances to potassium ions.

Primary afferent-evoked synaptic responses and slow potential generation in rat substantia gelatinosa neurons in vitro.

1. Primary afferent fiber-evoked synaptic responses and the mechanisms of spike and slow potential generation have been examined in adult rat substantia gelatinosa (SG) neurons in an in vitro transverse spinal cord slice preparation in which an attached dorsal root is retained. Intracellular recordings were made from SG neurons identified by morphological and electrophysiological criteria. Afferent fiber-evoked fast excitatory postsynaptic potentials (fast EPSPs) and slow EPSPs have been analyzed. 2. SG neurons had mean resting membrane potentials of -67.1 +/- 0.5 mV (mean +/- SE), mean input resistance of 257 +/- 17.7 (SE) M omega, and a mean time constant of 21.3 +/- 1.9 ms and exhibited spontaneous EPSPs. 3. Single low-intensity stimuli applied to the dorsal root using a suction electrode produced, in 70% of SG neurons, short-latency, presumed monosynaptic fast EPSPs which had a half decay time of 10-30 ms and an amplitude of 8-28 mV. The conduction velocity of afferent fibers evoking fast EPSPs was 2-7 m/s, corresponding to that of thinly myelinated A-delta-fibers. Dorsal root stimulation at higher intensities evoked, in 10% of SG neurons, long-latency and apparently monosynaptic EPSPs which had a time course and amplitude similar to that evoked by low-intensity stimulation. The conduction velocity of fibers evoking long-latency EPSPs was 0.4-2 m/s, suggesting that they constitute predominantly C-fibers. A-delta- and C-fiber-mediated fast EPSPs were detected in 20% of SG neurons examined. 4. Low-intensity stimuli produced slow EPSPs in 20% of SG neurons. Slow EPSPs were 3-15 mV in amplitude and of up to 2 min in duration. A-delta-fibers appeared to be responsible for the generation of slow EPSPs. Slow EPSPs were associated with an increase in membrane resistance and were decreased in amplitude with membrane hyperpolarization. 5. Action potentials in SG neurons had a mean amplitude of 76.3 +/- 1.1 mV and a mean duration of 1.0 +/- 0.07 ms. Na+ ions represent the main charge carrier during the rising phase of the action potential and Ca2+ ions contribute to the shoulder on the falling phase. 6. In 20% of SG neurons, subthreshold depolarizing pulses were followed by long-lasting slow-inactivating depolarizing potentials which were able to initiate spikes. The slow depolarizing potentials were blocked by TTX and enhanced by application of TEA and Ba2+, suggesting that Na+ and K+ are involved in this slow-inactivating potential.(ABSTRACT TRUNCATED AT 400 WORDS)

Extraglandular Innervation of the Salivary Cells of the Leech Haementeria Ghilianii: Neuronal Stimulation Elicits Gland-Cell Action Potentials and Secretion

ABSTRACT Each salivary gland cell of Haementeria extends a single process, or ductule, anteriorly into the proboscis; secretory products are released at the ductule ending. Some ductules secrete into the lumen of the proboscis and others at the outer surface of its tip, more than 5 cm from the gland in large leeches. Depolarization of a gland-cell body elicits action potentials which appear to be conducted along the ductule to its ending. Electrical stimulation of the proboscis tip elicits action potentials in those ductules which end there, and the impulses are propagated to the cell body (approx. 5cms−1). Bathing the salivary glands in calcium-free saline causes spontaneous repetitive firing in the cell bodies and also elicits secretion at the proboscis tip (bathed in normal saline); the action potential thus appears to be a stimulus for secretion. A paired stomatogastric nerve, from the brain, enters the proboscis near its base. Cobalt-filling of the nerve shows numerous cell bodies in the brain and first body ganglion, and an intricate network of fibres and a cluster of stained cell bodies near its entry point in the proboscis. Repetitive stimulation of the stomatogastric nerve produces action potentials in certain gland cells, after a delay of at least 15 s, and also elicits secretion. The action potentials are initiated near the ductule tip, and are conducted to the cell body. The salivary glands themselves do not appear to be innervated. Application of acetylcholine (ACh), dopamine or octopamine (10−4mol 1−1) does not initiate secretion. Neither dopamine nor octopamine excites the gland cells but ACh produces a transient suprathreshold depolarization of the cell body and occasionally elicits 1–3 ductule spikes when applied to the proboscis tip. 5-Hydroxytryptamine (5-HT) produces secretion when applied to the proboscis but not when applied to the glands alone; it does not excite the cells, indicating that the action potential is not the only stimulus for secretion. 5-HT produces a depolarization, and increase in membrane resistance, in the cell body, and prevents the rapid adaptation of action potentials which occurs during maintained depolarization. 7.Electrophoretic analysis shows that the protein compositions of secretions at the proboscis tip and in the lumen are completely different, with the tip apparently secreting only two major proteins. These same two protein bands occur in the cytoplasm of certain gland-cell bodies which can be distinguished in living glands on the basis of size and degree of staining with Methylene Blue. 8.Following stimulation of the stomatogastric nerve, secretory products at the proboscis tip can be seen to emerge from discrete points which appear to be single ductule endings. This presents the possibility of studying excitation-secretion coupling in single cells.

On the mechanism of barium induced diastolic depolarisation in isolated ventricular myocytes.

Barium can induce spontaneous activity in cardiac non-pacemaker cells. The mechanism of barium induced diastolic depolarisation was studied in isolated ventricular myocytes, using a microelectrode technique. Barium (0.05-0.2 mmol.litre-1) decreased resting potential and caused the membrane potential at the end of the action potential to undershoot the diminished resting value temporarily, thereby inducing diastolic depolarisation. Resting membrane resistance was increased by Ba but at the end of phase 3 repolarisation the resistance temporarily decreased below its steady state diastolic value. In presence of Ba, hyperpolarisation abolished or reversed diastolic depolarisation. At the end of phase 3 repolarisation, membrane resistance was decreased, whether diastolic depolarisation was present, absent or reversed. A high [K]o (15.4 mmol.litre-1) decreased Ba effects on action potential, membrane resistance and diastolic depolarisation. Caesium decreased the Ba induced diastolic depolarisation and the associated increase in membrane resistance, but had little effect on spontaneous activity at depolarised levels. Barium induced an oscillatory potential, with increased membrane resistance. Noradrenaline plus low [Ba]o, and high [Ba]o alone (1-5 mmol.litre-1), can induce spontaneous activity. Thus, in myocardial cells barium induces diastolic depolarisation at polarised levels by a voltage and time dependent block of potassium conductance, which is modulated by action potential voltage changes. However, as [Ba]o is increased, spontaneous activity at a depolarised level may be related to the decay of potassium currents and to oscillatory potentials.

Amiloride does not block taste transduction in the mouse (Slc:ICR)

1. 1. The receptor potential of the mouse taste cell was recorded with an intracellular microelectrode while taste stimuli were applied to the tongue surface of the anesthetized mouse. 2. 2. A membrane depolarization accompanied by an increase in membrane resistance was observed after a sucrose stimulus. 3. 3. A sodium-chloride stimulus initiated a membrane depolarization accompanied by a decrease in membrane resistance. 4. 4. Amiloride elicits a depolarization of the membrane and is accompanied by an increase in membrane resistance. 5. 5. Pre-adapting the tongue to amiloride, which is known as a potent sodium channel blocker, did not alter the responses to sodium-chloride and other taste stimuli.

The Effect of Diphenylamine-2-carboxylate on C1− Channel Conductance and on Excitability Characteristics of Rat Skeletal Muscle

The effect of diphenylamine-2-carboxylate (DPC), a blocker of the C1- conductive pathway in C1- transporting epithelia, has been evaluated in-vitro on the electrophysiological variables of rat extensor digitorum longus muscle fibres. DPC (5-240 microM) caused a dose-related increase of membrane resistance which was attributed entirely to a fall in C1- channel conductance (IC50, 120 microM), since potassium conductance was not affected by the treatment. DPC also modified fibre excitability. A significant dose-dependent increase was observed in the latency of the action potential and in the excitability of the membrane. DPC was less potent on striated fibres than anthracene-9-carboxylic acid, another specific blocker of C1- channel conductance. Moreover DPC was less potent on skeletal muscle than on C1- transporting epithelia. Morphological differences in the C1- channels or of the drug binding sites may account for the differences between tissues.

Membrane currents induced by pentylenetetrazol in identified neurons of Helix pomatia

After systemic application of pentylenetetrazol (PTZ), mammalian as well as molluscan neurons generate epileptic paroxysmal depolarization shifts. For a further analysis of these potential oscillations the membrane currents induced by local application of PTZ onto identified neurons of Helix pomatia were investigated. Different types of responses were obtained at membrane potentials negative and positive to ca. −30 mV. At holding potentials more negative than −30 mV, PTZ as a rule evoked an inward current, sometimes preceded by a brief outward current. In a few experiments only a solitary outward current was found. The amplitudes of the inward and outward currents increased towards more negative potentials. The inward current was associated with a decrease and the outward current with an increase in membrane resistance. Besides these findings pharmacological and ion substitution experiments indicate that the inward current represents an unspecific current. At holding potentials more positive than −30 mV, PTZ evoked a sequence of currents which was the same in all neurons. This stereotyped current sequence consisted of (i) an early inward current, (ii) an intermediate outward current, and (iii) a late long-lasting inward current. The amplitudes of all these components increased towards more positive potentials with the outward current being particularly enhanced. The early inward current and the following outward current were associated with a decrease and the late inward current with an increase of the membrane resistance. Besides these pharmacological and ion substitution experiments suggest that the early inward current represents a mixed sodium and calcium current. the intermediate outward current a calcium activated potassium current. The late inward current is assumed to be due to a decreased potassium conductance. On the basis of the present results, it may be concluded that the unspecific inward current in the negative potential range is involved in the initiation and the calcium dependent potassium current in the termination of spontaneously occuring paroxysmal depolarization shifts.

Direct effects of 3,4-dihydroxybutanoic acid γ-lactone and 2,4,5-trihydroxypentanoic acid γ-lactone on lateral and ventromedial hypothalamic neurons

The mechanism of central actions of endogenous sugar acids, 3,4-dihydroxybutanoic acid γ-lactone (3,4-DB) and 2,4,5-trihydroxypentanoic acid γ-lactone (2,4,5-TP) which have been newly identified as satiety and hunger substances respectively, was investigated. Intracellular recordings were made from neurons in the lateral hypothalamic area (LHA) of rats and ventromedial hypothalamic nucleus (VMH) of guinea pigs, brain referred to as feeding and satiety centers, respectively. The LHA neurons hyperpolarized by 3,4-DB show no change in membrane input resistance while the depolarized VMH neurons are associated with an increase in membrane resistance. The mechanisms related to the action of 3,4-DB on these hypothalamic neurons are similar to those in case of glucose on the glucose-sensitive neuron in the LHA and the glucoreceptor neuron in the VMH. 2,4,5-TP depolarized LHA neurons but hyperpolarized VMH neurons with a decrease in the membrane resistance. Our findings indicate that 3,4-DB and 2,4,5-TP have reciprocal effects on each LHA and VMH neurons, with regard to neuronal excitability.

Intracellular pH influences the resting membrane potential of isolated rat hepatocytes

This study in isolated rat hepatocytes sought to determine if the changes in membrane potential associated with intracellular alkalinization or acidification could be attributed to changes in K + conductance. Intracellular pH i was manipulated using the ‘NH 4 +-pulse’ method: inducing intracellular alkalinization with NH 4Cl (10 mM), and producing acidification by diluting the NH 4 +-loaded cells with ammonium ion-free buffer or by adding sodium proprionate. Membrane potential and resistance were measured in freshly isolated rat liver cells using intracellular microelectrodes. The results indicated that intracellular alkalinization was associated with hyperpolarization and decreased membrane resistance, whereas intracellular acidification caused depolarization with increased membrane resistance. As pH i-mediated electrogenic responses have been related to changes in K + conductance in other epithelial tissues, the influence of K + transport inhibitors on NH 4 +-evoked electrical effects was examined. NH 4Cl-evoked membrane potential changes were inhibited by the K + channel blockers, quinine and barium and in potassium depolarized cells (cells bathed in a high K + medium where [ K +] in = [ K +] out = 140mM). Furthermore, Rubidium-86 ( 36Rb +) efflux from preloaded hepatocytes, a measure of K + permeability, was enhanced following intracellular alkalinization but inhibited by intracellular acidification. Thus, these results indicate that pH i-evoked electrogenic effects in hepatocytes are mediated through changes in K + conductance.

Characterization and localization of a peripheral neural 5- hydroxytryptamine receptor subtype (5-HT1P) with a selective agonist, 3H-5-hydroxyindalpine

Peripheral neural 5-hydroxytryptamine (5-HT) receptors are different from both classes 5-HT1 and 5-HT2, which have been described from studies of 5-HT receptors in the brain. Recently, it has been shown that, as in the CNS, there is more than a single type of neural receptor for 5-HT in the enteric nervous system. One of these, called 5-HT1P, has a high affinity for 3H-5-HT, initiates a long-lasting depolarization of enteric neurons associated with an increase in membrane resistance, and is the physiological receptor through which enteric serotoninergic neurons mediate slow EPSPs. The other receptor, called 5-HT3 (5-HT2P), does not bind 3H-5-HT with high affinity, and initiates a brief depolarization of enteric neurons with decreased input resistance, but a physiological action of 5-HT mediated by these receptors has not yet been identified. Hydroxylated indalpines have been found to be agonists at 5-HT1P receptors. We have now examined 5-HT1P receptors using 5-hydroxyindalpine (5-OHIP) as a probe. The action of 5-OHIP on enteric neurons was determined electrophysiologically and compared with that of 5-HT; the binding of 3H-5-OHIP to isolated enteric membranes was studied by rapid filtration, and to frozen sections of tissue by radioautography. 3H-5-OHIP binding was compared with that of 3H-5-HT. 5-OHIP, like 5-HT, induced a triphasic response in most enteric neurons: an initial short-lived depolarization, during which input resistance fell, followed by recovery, and then a long-lasting depolarization, during which the input resistance increased. 5-OHIP bound saturably, reversibly, and with high affinity to enteric membranes (Kd = 7.6 +/- 0.7 nM; Bmax = 76 +/- 14 fmol/mg protein). Binding of 3H-5-OHIP was not inhibited by agents that bind to alpha- or beta-adrenoceptors, nicotinic or muscarinic receptors, histamine H1 or H2 receptors, or 5-HT1(A,B,C, or D), 5-HT2, or 5-HT3 receptors, but was displaced by substances, such as hydroxylated indoles and a dipeptide of 5-hydroxytryptophan (5-HTP-DP), that antagonize the binding of 3H-5-HT to enteric membranes or tissue sections. It is concluded that 5-OHIP is an agonist at peripheral neural 5-HT1P receptors and can be used to label these receptors selectively outside the brain. Radioautographs demonstrated enteric 5-HT1P receptors in the lamina propria of the intestinal mucosa and in the submucosal and myenteric plexuses. Extraenteric 5-HT1P receptors were also found in the skin and heart. It is suggested that 5-HT1P receptors may be found on subtypes of primary afferent nerve fibers.

In vitro effects of substance P on neonatal rat sympathetic preganglionic neurones.

1. Intracellular recordings were made from antidromically identified sympathetic preganglionic neurones (SPNs) in thin transverse neonatal rat thoracolumbar spinal cord slices. 2. Applied either by pressure ejection or superfusion, substance P (SP) caused a slow, monophasic depolarization in 60% of sympathetic preganglionic neurones; a biphasic response consisting of an initial hyperpolarization followed by a depolarization was observed in a few neurones. In addition, SP induced the occurrence of repetitive inhibitory postsynaptic potentials (IPSPs) in about 20% SPNs. 3. Low-Ca2+ or tetrodotoxin (TTX)-containing Krebs solution abolished the hyperpolarizing phase of the biphasic response and the small IPSPs, thereby augmenting the depolarizing response of SP. 4. SP-induced depolarizations were often associated with a moderate increase in membrane resistance. Generally, the response was made smaller on hyperpolarization and reversed at the membrane potential between -90 and -100 mV. These findings suggest that a reduction of membrane K+ conductance may underlie the depolarizing action of SP. 5. Subthreshold fast, excitatory postsynaptic potentials (EPSPs) evoked by stimulation of dorsal rootlets were consistently augmented during SP-induced depolarization, leading to cell discharge. 6. Focal stimulations elicited, in addition to a fast EPSP, a slow EPSP in about 40% of SPNs. The slow EPSP was often associated with an increased membrane resistance and became smaller on hyperpolarization. 7. In 15% of SPNs that generated a slow EPSP, the latter was reversibly abolished during SP-induced depolarization; the blockade persisted when the membrane potential was restored to the resting level by hyperpolarizing current. 8. It is concluded that SP is excitatory to SPNs and that its synaptic release may initiate a slow EPSP which serves to augment impulse transmission through SPNs. Further, it appears that inhibitory interneurones may also be sensitive to SP and their activation may provide a negative feed-back mechanism which can limit excessive excitation of SPNs by the peptide.