Purinergic fast excitatory postsynaptic potentials in myenteric neurons of guinea pig: distribution and pharmacology

BACKGROUND & AIMS: Adenosine triphosphate (ATP) acting at P2 receptors mediates some fast excitatory postsynaptic potentials (fEPSPs) in myenteric neurons of guinea pig ileum. The present studies investigate the distribution of purinergic fEPSPs along the length of the gut and characterize the P2-receptor subtype mediating fEPSPs. METHODS: Conventional intracellular electrophysiological methods were used to record from myenteric neurons in vitro. RESULTS: At a membrane potential of -97 +/- 1 mV, the amplitude (25 +/- 1 mV; n = 307) of fEPSPs was similar along the gut. Hexamethonium (100 micromol/L) inhibited fEPSPs in the gastric corpus by 98% +/- 1% (n = 31) and in the duodenum, ileum, taenia coli, proximal colon, and distal colon by 42%-55%. In the presence of hexamethonium, suramin (100 micromol/L) or the P2X antagonist pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid (PPADS, 10 micromol/L) reduced the control fEPSP amplitude in the duodenum, ileum, taenia coli, proximal colon, and distal colon by 71%-84%. The pharmacology of the purinergic fEPSPs was investigated in detail in the ileum. Noncholinergic fEPSPs were concentration-dependently (1-30 micromol/L) inhibited by PPADS (50%-inhibitory concentration, 3 micromol/L). In addition, alpha,beta-methylene 5'-adenosine triphosphate (1 micromol/L) also reduced purinergic fEPSPs. CONCLUSIONS: Fast EPSPs mediated in part through P2X receptors are prominent in myenteric neurons along the small and large intestines but are rare in the gastric corpus.     

Electrical activity of the proventriculus of the polychaete worm Syllis spongiphila

The straited myoepithelial cells of the proventriculus of Syllis spongiphila are composed of only one or two sarcomeres that may reach 40 mum in length. Experiments were performed to study some of their electrophysiological properties and their synaptic control. The mean resting potentials recorded in two different bathing media were 59-1 +/- 5-5 mV (S.D., n=91) and 62-5 +/- 6-3 mV (S.D., n=98). At rest the membrane potential is determined largely by permeability of the membrane to K+ ions, but the membrane is also permeable to other ions. On a semilogarithmic plot of membrane potential v. [K]o the mean slope of the data points from 9 to 90 mM-[K]o was 48 +/- 3 mV for a 10-fold change in [K]o. The anterior end of the animal was stimulated with a suction electrode to elicit activity of nerve fibres that innervate the proventriculus. Single indirect stimuli usually evoked hyperpolarizing or biphasic responses, and occasionally depolarizing responses, from the myoepithelial cells. The depolarizing synaptic potentials exhibited a faster time course than the hyperpolarizing ones. The rise time to peak ranged from 20 to 35 ms for simple depolarizations (n=32) and 25-75 ms for simple hyperpolarizations (n=103). Time to decay to half amplitude ranged from 20 to 55 ms for depolarizations (n=29) and 62-135 ms for hyperpolarizations (n=87). Low frequency (is less than or equal to 4 Hz) trains of indirectly applied stimuli elicited mainly hyperpolarizing responses; higher frequency (5-40 Hz) trains elicited complex responses composed of hyperpolarizations and depolarizations. Hyperpolarizations were selectively and reversibly abolished in chloride-free solutions. The reversal potential of the hyperpolarizing synaptic potential was -104 +/- 3 mV (S.D., n=8, 2 preparations). In calcium-free solution both hyperpolarizations and depolarizations were almost completely abolished. 4 mM-Mn2+ added to the bath almost completely abolished the depolarization but not the hyperpolarization. It was not clear whether Mn2+ acted at the presynaptic membrane, the postsynaptic membrane or both. The myoepithelial cells are electrically coupled. The mean space constant of five preparations was 0-52 mm (range 0-40-0-66 mm).     

NMDA receptor dependence of mGlu-mediated depression of synaptic transmission in the CA1 region of the rat hippocampus

1. The depression of synaptic transmission by the specific metabotropic glutamate receptor (mGlu) agonist (1S, 3R)-1-aminocyclopentane-1,3-dicarboxylate ((1S,3R)-ACPD) was investigated in area CA1 of the hippocampus of 4-10 week old rats, by use of grease-gap and intracellular recording techniques. 2. In the presence of 1 mM Mg2+, (1S,3R)-ACPD was a weak synaptic depressant. In contrast, in the absence of added Mg2+, (1S,3R)-ACPD was much more effective in depressing both the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) and N-methyl-D-aspartate (NMDA) receptor-mediated components of synaptic transmission. At 100 microM, (1S,3R)-ACPD depressed the slope of the field excitatory postsynaptic potential (e.p.s.p.) by 96 +/- 1% (mean +/- s.e.mean; n = 7) compared with 23 +/- 4% in 1 mM Mg(2+)-containing medium (n = 17). 3. The depressant action of 100 microM (1S,3R)-ACPD in Mg(2+)-free medium was reduced from 96 +/- 1 to 46 +/- 6% (n = 7) by the specific NMDA receptor antagonist (R)-2-amino-5-phosphonopentanoate (AP5; 100 microM). 4. Blocking both components of GABA receptor-mediated synaptic transmission with picrotoxin (50 microM) and CGP 55845A (1 microM) in the presence of 1 mM Mg2+ also enhanced the depressant action of (1S,3R)-ACPD (100 microM) from 29 +/- 5 to 67 +/- 6% (n = 6). 5. The actions of (1S,3R)-ACPD, recorded in Mg(2+)-free medium, were antagonized by the mGlu antagonist (+)-alpha-methyl-4-carboxyphenylglycine ((+)-MCPG). Thus, depressions induced by 30 microM (1S,3R)-ACPD were reversed from 48 +/- 4 to 8 +/- 6% (n = 4) by 1 mM (+)-MCPG. 6. In Mg(2+)-free medium, a group I mGlu agonist, (RS)-3, 5-dihydroxyphenylglycine (DHPG; 100 microM) depressed synaptic responses by 74 +/- 2% (n = 18). In contrast, neither the group II agonists ((2S,1'S,2'S)-2-(2'-carboxycyclopropyl)glycine; L-CCG-1; 10 microM; n = 4) and ((2S,1'R,2'R,3'R)-2-(2',3'-dicarboxycyclopropyl)glycine; DCG-IV; 100 nM; n = 3) nor the group III agonist ((S)-2-amino-4-phosphonobutanoic acid; L-AP4; 10 microM; n = 4) had any effect. 7. The depolarizing action of (1S,3R)-ACPD, recorded intracellularly, was similar in the presence and absence of Mg(2+)-AP5 did not affect the (1S,3R)-ACPD-induced depolarization in Mg(2+)-free medium. Thus, 50 microM (1S,3R)-ACPD induced depolarizations of 9 +/- 3 mV (n = 5), 10 +/- 2 mV (n = 4) and 8 +/- 2 mV (n = 5) in the three respective conditions. 8. On resetting the membrane potential in the presence of 50 microM (1S,3R)-ACPD to its initial level, the e.p.s.p. amplitude was enhanced by 8 +/- 3% in 1 mM Mg2+ (n = 5) compared with a depression of 37 +/- 11% in the absence of Mg2+ (n = 4). Addition of AP5 prevented the (1S,3R)-ACPD-induced depression of the e.p.s.p. (depression of 4 +/- 5% (n = 5)). 9. It is concluded that activation by group 1 mGlu agonists results in a depression of excitatory synaptic transmission in an NMDA receptor-dependent manner.     

The distal colon provides reserve storage capacity during colonic fluid overload

BACKGROUND: In addition to its absorptive function the capacity of the colon to retain fluid might be relevant in compensating for increased fluid loads and prevention of diarrhoea. The distal colon is considered to be mainly a conduit without extensive storage function. AIMS: To evaluate colonic volume capacity in a model of pure osmotic diarrhoea. METHODS: A non-absorbable, iso-osmotic solution (OS) containing polyethylene glycol (500 ml) was infused into the caecum of nine healthy volunteers; the control group (n = 5) received an equal amount of an easily absorbable electrolyte solution (ES). Fluids were radiolabelled with technetium-99m and gamma camera images were obtained for 48 hours. Counts in the proximal and distal colon were measured and regional and overall colonic transit and stool output were quantified. RESULTS: After OS, in contrast to ES, faecal output was increased significantly (p < 0.05), but colonic transit after OS was not different from transit after ES (p > 0.05). This indicates storage of OS in the colon: after OS infusion, counts in the proximal colon decreased linearly while the distal colon stored approximately 30% of radioactivity for the whole 48 hour study period. After OS, stool output correlated with distal (p < 0.01), but not with proximal (p > 0.05), colonic transit. In contrast, after ES, stool output was determined by proximal colonic transit (p < 0.05) but not by transit through the distal colon (p > 0.05). CONCLUSION: The distal colon retains non-absorbable fluid volumes extensively. In our model transit through the distal colon--but not the proximal colon--determined the time at which diarrhoea occurred.     

Tetanic stimulation induces short-term potentiation of inhibitory synaptic activity in the rostral nucleus of the solitary tract

Whole cell recordings from neurons in the rostral nucleus of the solitary tract (rNST) were made to explore the effect of high-frequency tetanic stimulation on inhibitory postsynaptic potentials (IPSPs). IPSPs were elicited in the rNST by local electrical stimulation after pharmacological blockade of excitatory synaptic transmission. Tetanic stimulation at frequencies of 10-30 Hz resulted in sustained hyperpolarizing IPSPs that had a mean amplitude of -68 mV. The hyperpolarization resulted in a decrease in neuronal input resistance and was blocked by the gamma-aminobutyric acid-A (GABAA) antagonist bicuculline. For most of the neurons (n = 87/102), tetanic stimulation resulted in a maximum hyperpolarization immediately after initiation of the tetanic stimulation, but for some neurons the maximum was achieved after three or more consecutive shock stimuli in the tetanic train of stimuli. When the extracellular Ca2+ concentration was reduced, the maximum IPSP amplitude was reached after several consecutive shock stimuli in the tetanic train for all neurons. Tetanic stimulation at frequencies of 30 Hz and higher resulted in IPSPs that were not sustained but decayed to a more positive level of hyperpolarization. In some neurons the decay was sufficient to become depolarizing and resulted in a biphasic IPSP. It was possible to evoke this biphasic IPSP in all the neurons tested if the cells were hyperpolarized to -75 to -85 mV. The ionic mechanism of the depolarizing IPSPs was examined and was found to be due to an elevation of the extracellular K+ concentration and accumulation of intracellular Cl-. Tetanic stimulation increased the mean 80-ms decay time constant of a single shock-evoked IPSP up to 8 s. The length of the IPSP decay time constant was dependent on the duration and frequency of the tetanic stimulation as well as the extracellular Ca2+ concentration. Afferent sensory input to the rNST consists of trains of relatively high-frequency spike discharges similar to the tetanic stimulation frequencies used to elicit the IPSPs in the brain slices. Thus the short-term changes in inhibitory synaptic activity in the slice preparation probably occur in vivo and may play a key role in taste processing by facilitating synaptic integration.     

Ammonia-induced depolarization of cultured rat cortical astrocytes

Exposure of cultured rat cortical astrocytes to increased concentrations of ammonia has been shown to induce morphological and biochemical changes similar to those found in hyperammonemic (e.g., hepatic) encephalopathy in vivo. Alterations of electrophysiological properties are not well investigated. In this study, we examined the effect of ammonia on the astrocyte membrane potential by means of perforated patch recordings. Exposure to millimolar concentrations of NH4Cl induced a slow dose-dependent and reversible depolarization. At steady state, i.e., after several tens of minutes, the cells were significantly depolarized from a resting membrane potential of -96.2 +/- 0.6 mV (n = 83, S.E.M.) to -89.1 +/- 1.6 mV (n = 7, S.E.M.) at 5 mM NH4Cl, -66.3 +/- 3.6 mV (n = 9, S.E.M.) at 10 mM NH4Cl and -50.4 +/- 2.5 mV (n = 12, S.E.M.) at 20 mM NH4Cl, respectively. In order to examine the underlying depolarizing mechanisms we determined changes in the fractional ion conductances for potassium, chloride and sodium induced by 20 mM NH4Cl. No significant changes were found in the fractional sodium or chloride conductances, but the dominating fractional potassium conductance decreased slightly from a calculated 0.86 +/- 0.04 to 0.77 +/- 0.04 (n = 9, S.E.M.). Correspondingly, we found a significant fractional ammonium ion (NH4+) conductance of 0.23 +/- 0.02 (n = 10, S.E.M.) which was blocked by the potassium channel blocker barium and, hence, most likely mediated by barium-sensitive potassium channels. Our data suggest that the sustained depolarization induced by NH4Cl depended on changes in intracellular ion concentrations rather than changes in ion conductances. Driven by the high membrane potential NH4+ accumulated intracellularly via a barium-sensitive potassium conductance. The concomitant decrease in the intracellular potassium concentration was primarily responsible for the observed slow depolarization.     

Isolated perfused rabbit colon crypts: stimulation of Cl- secretion by forskolin

The aim of this study was to characterize ion conductances and carrier mechanisms of isolated in vitro perfused rabbit colonic crypts. Crypts were isolated from rabbit colon mucosa and mounted on a pipette system which allowed controlled perfusion of the lumen. In non-stimulated conditions basolateral membrane voltage (Vbl) was -65 +/- 1 mV (n = 240). Bath Ba2+ (1 mmol/l) and verapamil (0.1 mmol/l) depolarized Vbl by 21 +/- 2 mV (n = 7) and 31 +/- 1 (n = 4), respectively. Lowering of bath Cl- concentration hyperpolarized Vbl from -69 +/- 3 to -75 +/- 3 mV (n = 9). Lowering of luminal Cl- concentration did not change Vbl. Basolateral application of loop diuretics (furosemide, piretanide, bumetanide) had no influence on Vbl in non-stimulated crypts. Forskolin (10(-6) mol/l) in the bath depolarized Vbl by 29 +/- 2 mV (n = 54) and decreased luminal membrane resistance. In one-third of the experiments a spontaneous partial repolarization of Vbl was seen in the presence of forskolin. During forskolin-induced depolarization basolateral application of loop diuretics hyperpolarized Vbl significantly and concentration dependently with a potency sequence of bumetanide > piretanide > or = furosemide. Lowering bath Cl- concentration hyperpolarized Vbl. Lowering of luminal Cl- concentration from 120 to 32 mmol/l during forskolin-induced depolarization led to a further depolarization of Vbl by 7 +/- 2 mV (n = 10). We conclude that Vbl of rabbit colonic crypt cells is dominated by a K+ conductance. Stimulation of the cells by forskolin opens a luminal Cl- conductance. Basolateral uptake of Cl- occurs via a basolateral Na+:2Cl-:K+ cotransport system.     

Oxytocin acts at V1 receptors to excite sympathetic preganglionic neurones in neonate rat spinal cord in vitro

Intracellular recordings were made from sympathetic preganglionic neurones (SPNs) in transverse slices of thoraco-lumbar spinal cord of young rats (12-20 days old). A small group of SPNs generally having higher membrane potentials (-70 mV) compared to a remaining group (-66 mV) showed spontaneous oscillations of their membrane potential. Oxytocin superfused in concentrations of 0.1-30 microM had four effects on SPNs, inducing slow depolarisation, EPSPs, IPSPs and brief rhythmic oscillations. The slow depolarisation was unaffected by TTX whereas this abolished the other changes. The oxytocin-induced depolarisation was associated with a slow inward current and was not reversed at membrane potentials negative to EK, it increased at more positive potentials and was still present in low Ca2+ and high Mg2+ solutions. These features of the oxytocin induced current are similar to those of the TTX resistant voltage dependent Na+ current described in brainstem autonomic neurones. Vasopressin superfused at concentrations of 0.1 microM to 30 microM had similar effects on SPNs to those of oxytocin. A comparison of the effects of oxytocin and vasopressin on the same neurones revealed that oxytocin was almost 10 times less potent than vasopressin. The effects of oxytocin were not mimicked by a selective oxytocin agonist but were mimicked by a selective vasopressin V1a agonist and blocked by a selective V1a antagonist. Therefore it is concluded that the effects of oxytocin on SPNs are mediated by the vasopressin V1a receptor. It is suggested that oxytocin and vasopressin terminals in the lateral horn are part of a descending system controlling oscillating networks of SPNs in the spinal cord.     

A comparison of the effects of selective metabotropic glutamate receptor agonists on synaptically evoked whole cell currents of rat spinal ventral horn neurones in vitro

1. Whole cell synaptic currents were recorded under voltage clamp from a total of 54 ventral horn neurones held near to their resting potential by the patch clamp technique in immature rat spinal cord preparations in vitro. Twenty eight neurones were identified, by antidromic invasion from ventral roots, as motoneurones. Excitatory postsynaptic currents (e.p.s.cs) of peak amplitude -480 pA +/- 66 s.e. mean and -829 +/- 124 pA were evoked respectively from the unidentified ventral horn neurones and the motoneurones in response to maximal activation of the segmental dorsal root. 2. The e.p.s.cs were depressed reversibly by the metabotropic glutamate agonists 1S3S-1-aminocyclopentane-1,3-dicarboxylate (1S3S-ACPD) (EC50 17.1 microM +/- 0.3 s.e. mean, n = 14) and L-2-amino-4-phosphonobutanoate (L-AP4) (EC50 = 2.19 +/- 0.19 microM, n = 15). Since both agonists independently produced more than 90% depression it is likely that the receptors that mediate their effects are present on the same presynaptic terminals. 3. When the Mg2+ concentration was raised from 0.75 mM to 2.75 mM together with the addition of 50 microM D-2-amino-5-phosphonopentanoate (AP5), a treatment which would increase the proportion of monosynaptic component in the e.p.s.c. the concentration-effect plots for both 1S3S-ACPD (EC50 1.95 +/- 0.4 microM, n = 8) and L-AP4 (EC50 0.55 +/- 0.20 microM, n = 7) were shifted to the left, suggesting that monosynaptic e.p.cs of primary afferents to ventral horn neurones are more susceptible to L-AP4 and 1S3S-ACPD than are other synapses in polysynaptic pathways. 4. lS3S-ACPD (20 and 50 microM) also caused mean sustained inward currents of 95 +/- 31 pA (n = 6) and248 +/- 49 pA (n = 10) respectively. In the combined presence of AP5 (50 microM) and Mg2+ (2.75 mM) themean response to 50 microM lS3S-ACPD was reduced to 106+/- 18 pA (n = 4). In the presence of tetrodotoxin(1 microM) the corresponding value was 48 +/- 6 pA (n = 4). Similar sustained inward currents produced by N-methyl-D-aspartate (NMDA) were almost abolished to < 10 pA in the presence of AP5 and 2.75 mMMg2+. In the presence of tetrodotoxin the maximum inward current produced by NMDA was undiminished. Thus a large component of the excitatory action of lS3S-ACPD was mediated at non-NMDA receptors both directly at the patch-clamped neurones and indirectly by synaptic relay.     

Development of a generalized phenomenological model describing the kinetics of the enzymatic hydrolysis of NaOH-treated pine wood

It was assumed that the enlarged caecum and the accumulation of semiliquid contents in germfree rats is accompanied by changes in sodium absorption. Transepithelial sodium fluxes were studied under Ussing chamber conditions across epithelial sheets of the caecum and colon of germfree (GF) and specific-pathogen free (SPF) rats. Net sodium transport was highest in the proximal colon and in the proximal segment of the distal colon; it was considerably lower in the caecum and in the distal segment of the distal colon. In the caecum and proximal colon of the GF rats, the electroneutral sodium absorption was increased as compared to the SPF rats. In the proximal segments of the distal colon, no differences were seen. In the distal segment of the distal colon, the mainly electroneutral sodium transport in the SPF rats was changed into electrogenic transport in the GF rats. These differences may be due to the increased aldosterone levels of the GF rats.     

Voltage- and use-dependent inhibition of Na+ channels in rat sensory neurones by 4030W92, a new antihyperalgesic agent

1. Whole cell patch clamp techniques were used to study the effects of 4030W92 (2,4-diamino-5-(2,3-dichlorophenyl)-6-fluoromethylpyrimidine), a new antihyperalgesic agent, on rat dorsal root ganglion (DRG) neurones. 2. In small diameter, presumably nociceptive DRG neurones under voltage-clamp, 4030W92 (1-100 microM) produced a concentration-related inhibition of slow tetrodotoxin-resistant Na+ currents (TTXR). From a holding potential (Vh) of -90 mV, currents evoked by test pulses to 0 mV were inhibited by 4030W92 with a mean IC50 value of approximately 103 microM. 3. The inhibitory effect of 4030W92 on TTX(R) was both voltage- and use-dependent. Currents evoked from a Vh of -60 mV were inhibited by 4030W92 with a mean IC50 value of 22 microM, which was 5 fold less than the value obtained at -90 mV. Repeated activation of TTX(R) by a train of depolarizing pulses (5 Hz, 20 ms duration) enhanced the inhibitory effects of 4030W92. These data could be explained by a preferential interaction of the drug with inactivation states of the channel. In support of this hypothesis 4030W92 (30 microM) produced a significant hyperpolarizing shift of 10 mV in the slow inactivation curve for TTX(R) and markedly slowed the recovery from channel inactivation. 4. Fast TTX-sensitive Na+ currents (TTXs) were also inhibited by 4030W92 in a voltage-dependent manner. The IC50 values obtained from Vhs of -90 mV and -70 mV were 37 microM and 5 microM, respectively. 4030W92 (30 microM) produced a 13 mV hyperpolarizing shift in the steady-state inactivation curve of TTXs. 5. High threshold voltage-gated Ca2+ currents were only weakly inhibited by 4030W92. The reduction in peak Ca2+ current amplitude produced by 100 microM 4030W92 was 20+/-6% (n=6). Low threshold T-type Ca2+ currents were inhibited by 17+/-8% and 43+/-3% by concentrations of 4030W92 of 30 microM and 100 microM, respectively (n=6). 6. Under current clamp, some cells exhibited broad TTX-resistant action potentials whilst others showed fast TTX-sensitive action potentials in response to a depolarizing current injection. In most cells a long duration (800 ms) supramaximal current injection evoked a train of action potentials. 4030W92 (10-30 microM) had little effect on the first spike in the train but produced a concentration-related inhibition of the later spikes. The number of spikes per train was significantly reduced from 9.7+/-1.5 to 4.2+/-1.0 and 2.6+/-1.1 in the presence of 10 microM and 30 microM 4030W92, respectively (n=5). 7. Thus, 4030W92 is a potent voltage- and use-dependent inhibitor of Na+ channels in sensory neurones. This profile can be explained by a preferential action of the drug on a slow inactivation state of the channel that results in a delayed recovery to the resting state. This state-dependent modulation by 4030W92 of Na+ channels that are important in sensory neurone function may underlie or contribute to the antihyperalgesic profile of this compound observed in vivo.     

Neuronal pathways for activation of inhibitory interneurons in pyriform cortex of the rabbit

The neuronal pathways responsible for the fast inhibitory postsynaptic potentials (IPSPs) elicited in principal cells in the pyriform cortex (PC) by volleys from the olfactory bulb (OB), the lateral olfactory tract (LOT), the anterior commissure (AC), and the deep-lying structures of the PC (DPC) were studied in the rabbit. The central latencies of the fast IPSPs (measured from the onset of the monosynaptic excitatory postsynaptic potential (EPSP) elicited by volleys through the LOT) ranged between 3.0 and 9.3 ms (5.5 +/- 1.3 (SD) ms; n = 54) in the case of OB shocks and between 4.5 and 6.5 ms (5.1 +/- 0.7 (SD) ms; n = 7) in the case of LOT shocks. The onset latencies of the fast IPSPs were between 2.5 and 11.8 ms (5.1 +/- 1.8 (SD) ms; n = 66) in the case of DPC shocks and between 3.5 and 10.1 ms (5.8 +/- 1.5 (SD) ms; n = 61) in the case of AC shocks. The conditioning OB or LOT shocks almost completely eliminated the LOT-evoked fast IPSP when the testing shock was applied at the peak period of the conditioning slow IPSP. The conditioning OB shocks also eliminated the initial part of the OB-evoked fast IPSP, leaving the later part of the fast IPSP almost unchanged. Thus, the onset latency of the OB-evoked fast IPSP was lengthened by 7.1 +/- 2.9 (SD) ms (n = 35) by the conditioning OB shock. The conditioning OB or DPC shocks left the peak amplitude of the DPC-evoked fast IPSP almost unaffected. Similarly, the conditioning OB or AC shocks left the peak amplitude of the AC-evoked fast IPSP almost unaffected. The conditioning OB, DPC, or AC shocks had only a slight influence on the onset latency of the DPC- or AC-evoked fast IPSPs. Rhythmical steps at intervals of 3-5 ms were observed in the rising phase of the OB-evoked fast IPSP. This was interpreted as a result of a repetitive impingement of interneuronal discharges on the impaled cells. Spatial facilitation was observed among the fast IPSPs evoked by volleys from the OB, DPC, and AC when shocks were applied at suitable intervals. A slight facilitation was also seen between the LOT-evoked fast IPSP and the DPC- or AC-evoked fast IPSP. These results were interpreted as a result of the convergence of excitatory synaptic inputs onto the presumed inhibitory interneurons from the four structures of the brain. A temporal facilitation of the fast IPSPs was observed when the OB, DPC, or AC shocks were applied repetitively at short intervals. This suggests a temporal facilitation of the spike discharges of the presumed inhibitory interneurons under similar conditions. From these results, criteria were determined for identifying the inhibitory interneurons.     

The effects of catechol on various membrane conductances in lumbar sympathetic postganglionic neurones of the guinea-pig

The effects of catechol on membrane properties in lumbar sympathetic postganglionic neurones isolated from guinea-pigs were studied in vitro in current and voltage clamp using single intracellular microelectrodes. Neurones with properties characteristic of two previously described classes of neurone (phasic and tonic) were studied. Catechol (3-12 mM) produced a few mV depolarization and a dose-dependent increase in membrane resistance which were both larger in tonic than in phasic neurones. In the presence of catechol, both phasic and tonic neurones fired only a single action potential at the beginning of a maintained depolarizing current step. In both neurone types, catechol reduced action potential amplitude and slowed its time course. The peak of the afterhyperpolarization became delayed and reduced in amplitude, particularly in tonic neurones. The time constant of inactivation of IA was reduced by catechol without change in the voltage sensitivity of activation or inactivation: IC50 was 3 mM in phasic and 4 mM in tonic neurones. Catechol also blocked a slow voltage-activated K+ current (resembling ID) that was present in many tonic neurones. Catechol did not modify the slow calcium-activated potassium current (gKCa1) or the anomalous rectifier; neither did it appear to affect the fast calcium-activated potassium current (IC) or the delayed rectifier. Catechol did not change the overall rate of spontaneous synaptic activity nor enhance the release of quanta of ACh from preganglionic terminals evoked by nerve stimulation. We conclude that, in addition to blocking IA, catechol blocks the slow ID-like current in sympathetic neurones. It also has a profound effect on the action potential probably by increasing inactivation of voltage-dependent Na+ channels. The change from tonic to phasic discharge in tonic neurones cannot be attributed solely to its effects on IA.     

Evaluation of driving performance in patients with juvenile macular dystrophies

The aim of the present study was to search for electrophysiological effects of human lipoproteins on membrane currents in mouse peritoneal macrophages which had been cultured for 5 to 20 days. Whole-cell currents were recorded by using a voltage-clamp technique. Low density lipoprotein (LDL, 100 micrograms/ml) increased a slowly activating nonspecific cation current (iso) in the positive potential range to 244 +/- 23% of the reference (test potential + 55 mV, n = 13, P < 0.005). Augmentation of current resulted out of a negative shift of the activation curve along the voltage axis (-22 mV) and an increase of maximally available current. Furthermore, LDL increased a rapidly activating outward current (ifo) at test potentials positive to the potassium equilibrium potential. At +55 mV ifo-amplitude increased to 165 +/- 14% of reference (n = 16, P < 0.005). LDL-induced effects on ifo-current could be mimicked by application of the calcium ionophore A 23,187 (1 mumol/l) which led to an increase of ifo-current to 161 +/- 25% of the reference (test potential +55 mV, n = 11, P < 0.005). Acetylated-LDL (100 micrograms/ml, 5-15 min) produced no significant effect on the membrane currents under investigation.     

Inhibition of calcium currents in rat colon sensory neurons by K- but not mu- or delta-opioids

Inhibition of calcium currents in rat colon sensory neurons by kappa- but not mu- or delta-opioids. J. Neurophysiol. 80: 3112-3119, 1998. We previously reported that kappa-, but not mu- or delta-opioid receptor agonists (ORAs) have selective, potentially useful peripheral analgesic effects in visceral pain. To evaluate one potential site and mechanism by which these effects are produced, we studied opioid effects on high-voltage activated (HVA) Ca2+ currents in identified (Di-I) pelvic nerve sensory neurons from the S1 dorsal root ganglion (DRG). Results were compared with opioid effects on cutaneous neurons from L5 or L6 DRG. Di-I-labeled DRG cells were voltage clamped (perforated whole cell patch clamp), and HVA Ca2+ currents were evoked by depolarizing 240-ms test pulses to +10 mV from a holding potential of -60 mV. Neither mu-ORAs (morphine, 10(-6 )M, n = 16; [D-Ala2, N-Me-Phe4, Gly-ol5] enkephalin, 10(-6 )M, n = 12) nor delta-ORAs ([D-Pen2, D-Pen5] enkephalin, 10(-7 )M, n = 16; SNC-80, 10(-7 )M, n = 7) affected HVA Ca2+ currents in colon sensory neurons. In contrast, the kappa-ORAs U50, 488 (10(-6 )M), bremazocine (10(-6)M), and nalBzoH (10(-6 )M) significantly attenuated HVA Ca2+ currents in colon sensory neurons; effects on cutaneous sensory neurons were variable. A nonreceptor selective concentration of naloxone (10(-5 )M) and nor-BNI (10(-6 )M), a selective kappa-opioid receptor antagonist, reversed the inhibitory effect of kappa-ORAs. In the presence of N-, P-, or Q-, but not L-type Ca2+ channel antagonists, the effect of U50,488 on HVA Ca2+ currents was significantly reduced. Pretreatment with pertussis toxin (PTX) prevented the inhibition by U50,488. These results suggest that kappa-opioid receptors are coupled to multiple HVA Ca2+ channels in colon sensory neurons by a PTX-sensitive G protein pathway. We conclude that inhibition of Ca2+ channel function likely contributes in part to the peripheral analgesic action of kappa-ORAs in visceral nociception.     

Actions of cisplatin on the electrophysiological properties of cultured dorsal root ganglion neurones from neonatal rats

In this study we have employed the whole cell patch clamp technique to investigate the effects of an anti-cancer drug cisplatin on basic electrophysiological properties of cultured dorsal root ganglion neurones from neonatal rats. The results show that within the clinical concentration range, cisplatin (0.1 to 10 microM) caused a decrease in input conductance, and complex changes in resting membrane potential in these cultured sensory neurones. The dominant effects of cisplatin on input conductance may be due to inhibition of leak conductances. Transplatin (5 microM) was significantly less effective than cisplatin at reducing input conductance which suggests a degree of stereoselectivity. Cisplatin (1 to 5 microM) transiently increased excitability of the cultured neurones as reflected by a reduction in the threshold for activation of action potentials by 8 mV. The rise time, peak amplitude and duration of action potentials were not changed by acute application of 5 microM cisplatin. Long term treatment of neurones with cisplatin (5 microM), for up to 1 week reduced the viability of the cultures, and attenuated neurone excitability, although input conductance of the cells was significantly increased to 322 +/- 49 M omega (n = 9) compared with controls of 210 +/- 20 M omega (n = 30; P < 0.05). Acute and chronic treatment of cultured neurones with cisplatin therefore produced contrasting actions.     

Glucose kinetics during exercise in trained men

The effects of a novel bradycardic agent Zeneca ZM 227189 (4-(N-ethyl-N-phenylamino)-1,2-dimethyl-6-(methylamino) triazinium iodide) were tested on the inward rectifying properties of guinea-pig substantia nigra pas compacta (SNC) and guinea-pig olfactory cortical cells recorded in vitro. In SNC neurones, ZM 227189 (10-100 microM) produced a dose-dependent block of the slow anomalous rectifier; under voltage clamp, a clear reduction was seen in the amplitude of the slow inward current (Ih) relaxation evoked by negative voltage commands from a holding potential of -60 mV. ZM 227189 (50-100 microM) induced an irreversible block of the Ih current after 10-15 min exposure. A similar block of Ih was observed following application of 5 mM Cs+. ZM 227189 had little effect on other membrane properties. By contrast, in olfactory cortical neurones, ZM 227189 (100 microM) induced an increase in the input resistance (approximately 20%) and cell excitability, accompanied by a small (< 2 mV) hyperpolarization; these effects were also not reversible. Activation of the fast (K(+)-mediated) inward rectifier at negative membrane potentials remained unaffected. Lower concentrations (1-10 microM) of ZM 227189 had no obvious effect on cortical cell properties. Our data indicate that ZM 227189 is a potent and apparently selective blocker of Ih in substantia nigra neurones, but has no effect on the fast-type inward rectifier in olfactory cortical cells.     

Structural and functional organization of the enteric nervous system in the stomach

This report summarises some features of the gastric enteric nervous system in the guinea-pig model. Particular attention has been paid to relations between neurochemical properties, electrophysiological and putative function of enteric neurones. (1) Cholinergic and nitrergic neurones form separate neuronal populations. (2) Ascending neurones outnumbered descending ones. (3) Transmitter-phenotype and projection were related: cholinergic neurones were primarily ascending while nitrergic neurones were mainly descending. (4) The neurochemical code, i.e. the transmitter colocalisation, could be related to the function of enteric neurones. Colocalisation of substance P and/or enkephaline in cholinergic neurones was characteristic for ascending excitatory muscle neurones. Descending inhibitory muscle neurones were nitrergic often colocalising the neuropeptides neuropeptide Y (NPY) and vasoactive intestinal peptide (VIP). In the intrinsic innervation of the gastric mucosa NPY/VIP was abundant and colocalised in ascending cholinergic as well as descending nitrergic neurones. (5) The vast majority of ascending and descending interneurones were cholinergic and often colocalised NPY. (6) The majority of descending mucosa and descending inhibitory muscle neurones were tonically-firing neurones. Our results revealed the characteristics of some neural components within the enteric nervous system of the stomach which are involved in modulation of mucosa and muscle functions. It may be concluded that muscle and mucosa functions are under the control of the enteric nervous system which contains distinct populations responsible for motor and secretory activity.     

Characterization of mechanosensitive pelvic nerve afferent fibers innervating the colon of the rat

1. Single-unit activity was recorded from S1 sacral dorsal root afferent fibers in the anesthetized rat. A total of 364 afferent fibers were identified by electrical stimulation of the pelvic nerve and subsequently tested for response to colorectal distension (CRD) and urinary bladder distension (UBD). Sixty-seven percent (n = 244) of the fibers were unmyelinated C-fibers and 33% (n = 120) were thinly myelinated A delta-fibers. 2. In three initial experiments, 35 fibers were identified by pelvic nerve stimulation and tested for response only to CRD; none of these fibers responded to CRD. In 20 subsequent experiments, 329 pelvic nerve afferent fibers were tested for response to CRD and UBD. Thirty-four percent (n = 112) of the 329 fibers were unresponsive to noxious CRD (80 mmHg) or to UBD (slow filling < or = 100 mmHg), 44% (n = 146) responded to UBD, 16% (n = 53) responded to CRD, and 6% (n = 18) responded to mechanical stimulation of the anal mucosa. 3. Of the total of 53 pelvic nerve afferent fibers that responded to CRD, 43 (81%) were C-fibers (mean: 1.5 m/s) and 10 (19%) were A delta-fibers (mean: 4.7 m/s). Fifteen of the CRD-sensitive fibers had no resting activity, whereas 38 fibers exhibited some resting activity (mean: 2.6 imp/s). 4. Reproducibility of responses to repeated CRD (80 mmHg, 30s, 10 trials at 4-min intervals) was tested in 17 fibers. In 16, responses to repeated distension were reproducible without evidence of facilitation or inhibition of subsequent responses. One fiber gave greater responses during the 9th and 10th trials. 5. Responses to graded CRD were studied in 44 fibers. All fibers exhibited monotonic, increasing stimulus-response functions < or = 80 mmHg of distension. Thresholds for response of the 44 fibers were determined after extrapolation of the least-squares linear-regression line to the ordinate and varied between 0 and 40 mmHg. Two populations of pelvic nerve afferent fibers in the colon were apparent: low threshold (LT) afferent fibers had a mean threshold of 2.9 mmHg (range: 0-10 mmHg; n = 34) and high threshold (HT) afferent fibers had a mean threshold of 32.6 mmHg (range: 28.0-40.0 mmHg; n = 10). 6. Chemosensitivity to bradykinin (BK) was tested in nine LT fibers. Seven fibers responded to BK (0.1 to 100 micrograms/kg ia) and two fibers did not respond up to 100 micrograms/kg of BK. Responses to BK tested in three fibers were dose dependent.(ABSTRACT TRUNCATED AT 400 WORDS)     

Acute pseudo-obstruction of the colon (Ogilvie's syndrome): advances in management

The management of the patients with acute pseudo-obstruction of the colon (APOC) still represents a matter of debate. To better evaluate and compare the effectiveness of various therapeutic approaches in the management of APOC 29 patients were considered. These were included according to three consecutive periods in: group A (1977-1982) concerning patients who underwent medical treatment alone (n = 8) or endoscopic (n = 4) and surgical (n = 1) decompression; group B (1983-1990) in which the management was based on simple endoscopic decompression (n = 10); group C (1991-1995) including patients in whom placement, under fluoroscopic control, of a tube in the cecum following endoscopic decompression was provided (n = 6). Mean time required for resolution of colonic distension was 2.3 (+/- 0.50 SD) days in patients who underwent endoscopic decompression and tube placement, as compared to 4.5 (+/- 2.47 SD) days in the group of patients treated either with conservative measures or simple endoscopic decompression (p = 0.04). No recurrence occurred after colonoscopic decompression and tube placement while colonic distension recurred in 4 of 14 patients managed by simple endoscopic decompression (0% vs. 28.6%, n.s.). Our experience showed that endoscopic decompression is an effective method, moreover if associated with the placement of an indwelling tube into the right colon. This method, for its easiness and safeness, besides its effectiveness in preventing the recurrence of colonic distension, may be surely considered an advance in the management of acute pseudo-obstruction of the colon.