Prolyl-leucyl-glycinamide (PLG) facilitates morphine dependence.

The development of physical dependence to morphine is facilitated in rats treated with [desglycinamide (9), arginine (8)] -vasopressin (DG-AVP) or with oxytocin. Oxytocin appeared to be more potent than DG-AVP. The essential elements of vasopressin and oxytocin required for facilitating morphine tolerance and physical dependence, are located in the C-terminal part of these hormones. It was found that the C-terminal tripeptide of oxytocin, prolylleucyl-glycinamide, was the most potent oligopeptide in this respect.     

Clotrimazole-sensitive K+ currents regulate pacemaker activity in interstitial cells of Cajal

Interstitial cells of Cajal (ICC) are pacemaker cells for gut peristaltic motor activity. Compared with cardiac pacemaker cells, little is known about mechanisms that regulate ICC excitability. The objective of the present study was to investigate a potential role for clotrimazole (CTL)-sensitive K currents (I(CTL)) in the regulation of ICC excitability and pacemaker activity. ICC were studied in situ and in short-term culture by using the whole cell patch-clamp configuration. In situ, ICC exhibited spontaneous transient inward currents followed by transient outward currents. CTL blocked outward currents, thereby increasing the net inward currents, and depolarized ICC, thereby establishing CTL-sensitive channels as regulators of ICC pacemaker activity. In short-term culture, a I(CTL) was identified that showed increased conductance when depolarized from the resting membrane potential to 0 mV and subsequent inward rectification at further depolarized potentials. The I(CTL) markedly increased with increasing intracellular calcium and was insensitive to the ether-à-go-go-related K channel blocker E-4031 and the large-conductance calcium-activated K channel blocker iberiotoxin. I(CTL) contributed 3-9 nS to the whole cell conductance at 0 mV membrane potential under physiological conditions; it was fast activating (tau = 88 ms), showed little time-dependent inactivation, and exhibited a deactivation time constant of 38 ms. The nitric oxide donor sodium nitroprusside (SNP) increased I(CTL). Single-channel activity, activated by calcium and SNP, was inhibited by CTL, with a single-channel conductance of approximately 38 pS. In summary, ICC generate a I(CTL) on depolarization through an intermediate-conductance calcium-activated K channel that regulates pacemaker activity and ICC excitability.     

The noradrenaline and serotonin content in symmetrical parts of the normal rat brain and during learning and peptide administration]

Content was compared of noradrenaline (NA) and serotonine (5-OT) in the right and left halves of the rats brain in norm, at elaboration of defensive conditioned reflexes of two-ways avoidance (CRTWA) and at administration of neuropeptides influencing the learning and memory--dezglycilargininvasopressin (DG-AVP), ACTH4-7 pro-gli-pro and dalargin. The conducted studies showed that in control animals the content of NA in the cortex of the right hemisphere was significantly higher than in the cortex of the left one. For the content of 5-OT in symmetric brain parts no significant differences were revealed. Under the elaboration of CRTWA the asymmetry of NA content was not eliminated. Systemic administration of DG-AVP, ACTG4-7 pro-gli-pro and dalargin practically did not change the content of 5-OT, but reduced the content of NA in the cortex and the rest of the brain, and the content of NA in the right and left cortex was equalized. The obtained data point to the asymmetric character of neuropeptides action and to greater resistance of 5-OT-ergic brain system to functional load and to administration of peptides in comparison with NA-ergic system.     

A comparative analysis of the effect of an analog of vasopressin on functionally different neurons in the grape snail

Application of desglycine-argininvasopressin (DG-AVP) differently influenced different types of cells of snail isolated central nervous system. In neurosecretory cells an increase of spontaneous impulse activity took place and, as a rule, bursts of impulses appeared, most often of synaptic origin, excluding PPa1 neurones and one of the neurosecretory cells of the left parietal ganglion. The increase of the bursts activity in these cells was based on the increase of the amplitude of membrane potential waves. Under the influence of neurosecretory cells system activation, EPSPs frequency and amplitude in secondary-sensory neurones increased, which led to a greater probability of the action potentials appearance. At prolonged action the spontaneous EPSPs in these cells began to group in bursts. Excitability and membrane resistance of these cells remained unchanged. DG-AVP had no influence on primary-sensory neurones and motoneurones.     

Resting membrane potentials of pacemaker and non pacemaker areas in rat uterus

Microelectrode studies of pacemaker and non pacemaker cells in pregnant rat uterus have shown the pacemaker areas to have a constant value of RMP throughout pregnancy which was always significantly smaller than that of non pacemaker cells. The development of new pacemaker areas was associated with membrane depolarization. A number of agents and conditions which caused membrane depolarization also induced pacemaker activity in previously non pacemaker areas but did so at different levels of membrane depolarization. Potassium depolarization failed to induce pacemaker activity. It is concluded that a low level of RMP is an important factor but not sufficient alone to explain the occurence of pacemaker activity.The resting membrane potentials (RMP) of spontaneously active smooth muscles are appreciably smaller than those of nonspontaneously active muscles (3) and comparable in magnitude to those of other tissues showing spontaneous activity such as the frog sinus venosus(7), rabbit sino-auricular node (10) and embryonic heart muscle (6). In intestinal smooth muscle, where all cells appear to be spontaneously active, a clear relationship can be demonstrated between fluctuating levels of RMP and the incidence of action potential activity, and the ionic and metabolic basis of slow wave activity has been extensively investigated (5, 8). In other smooth muscles, such as the ureter and uterus, where electrical activity arises from pacemaker areas (11, 12), the underlying causes of spontaneous activity are less well understood and the relationships between pacemaker activity, RMP and excitability have not been clearly defined. As an initial approach to studying this problem we have chosen to investigate the relationship between RMP and pacemaker activity in the uterus of the pregnant rat.     

Dynamics of changes in the excitability of Helix lucorum neurons in response to an analog of vasopressin

Application of desglycine-arginine-vasopressin to spontaneously nonactive command neurones of the snail's defensive reflex in the process of low-frequency (2-4 min intervals) intracellular stimulation led under certain conditions to an increase of excitability that was expressed in the shortening of latency of action potential generation, increase of the number of action potentials in response to a stimulus of fixed value, increase of membrane resistance, lowering of critical level of membrane depolarization and amplifying of pacemaker activity. However in spite of unidirectional changes of all the recorded parameters, the increases of values, opposite to the latency, did not correlate with the increases of membrane resistance and correlated well with the changes of depolarization critical level. If desglycine arginine-vasopressin was added to the medium during worsening of the neurones' excitability was probably caused by the influence of desglycine-arginine-vasopressin on the membrane active properties.     

Effect of a vasopressin analog on the chemoreactive properties of sensorimotor cortex neurons

Subcutaneous injection of 10 micrograms desglycilargininvasopressin (DG-AVP) does not alter the mean frequency of background unit activity of sensorimotor cortical neurons. However, the pattern of impulse activity is essentially changed. At the same time the reactions of sensorimotor cortical neurons to microiontophoretic administration of acetylcholine and noradrenaline experience definite changes. It is suggested that the DG-AVP-induced changes in chemoreactive properties of neurons underlie the effect of this peptide on the learning and memory.     

The Strategies of Behavioral Control in a Coelenterate

Although the cellular substrates of behavioral coordinationare uncertain in the hydroid Tubularia, much is known aboutthe strategies of behavioral control. The picture which emergesis that of an animal with diffusely distributed sites capableof initiating spontaneous activity, regional coordination ofpotential pacemaker loci to form pacemaker systems, and a loosehierarchical organization of pacemaker systems. At least onepacemaker system shows a short term increase in excitabilityand a longer duration depression of excitability following firing.The short-term excitability increase gives a tendency to firein bursts, the long-term depression acts as an intrinsic inhibitoryfeedback to terminate bursts and to control output frequency.All the known interactions between pacemaker systems are excitatory.Exogenous stimuli can excite a conducting system which inhibitsmost of the pacemaker systems, and one pacemaker system specificallyinhibits one set of muscles.     

Distribution of Kv1-like potassium channels in the electromotor and electrosensory systems of the weakly electric fish Apteronotus leptorhynchus

The electromotor and electrosensory systems of the weakly electric fish Apteronotus leptorhynchus are model systems for studying mechanisms of high-frequency motor pattern generation and sensory processing. Voltage-dependent ionic currents, including low-threshold potassium currents, influence excitability of neurons in these circuits and thereby regulate motor output and sensory filtering. Although Kv1-like potassium channels are likely to carry low-threshold potassium currents in electromotor and electrosensory neurons, the distribution of Kv1 alpha subunits in A. leptorhynchus is unknown. In this study, we used immunohistochemistry with six different antibodies raised against specific mammalian Kv1 alpha subunits (Kv1.1-Kv1.6) to characterize the distribution of Kv1-like channels in electromotor and electrosensory structures. Each Kv1 antibody labeled a distinct subset of neurons, fibers, and/or dendrites in electromotor and electrosensory nuclei. Kv1-like immunoreactivity in the electrosensory lateral line lobe (ELL) and pacemaker nucleus are particularly relevant in light of previous studies suggesting that potassium currents carried by Kv1 channels regulate neuronal excitability in these regions. Immunoreactivity of pyramidal cells in the ELL with several Kv1 antibodies is consistent with Kv1 channels carrying low-threshold outward currents that regulate spike waveform in these cells (Fernandez et al., J Neurosci 2005;25:363-371). Similarly, Kv1-like immunoreactivity in the pacemaker nucleus is consistent with a role of Kv1 channels in spontaneous high-frequency firing in pacemaker neurons. Robust Kv1-like immunoreactivity in several other structures, including the dorsal torus semicircularis, tuberous electroreceptors, and the electric organ, indicates that Kv1 channels are broadly expressed and are likely to contribute significantly to generating the electric organ discharge and processing electrosensory inputs.     

The paradoxically high reactivity of septal neurons in hibernating ground squirrels to endogenous neuropeptides is lost after chronic deafferentation of the septum from the preopticohypothalamic areas

The effect of neuropeptides (TSKYR, TSKY and DY) and neurotransmitters (serotonin and noradrenaline) on the activity of medial septum (MS) neurons from the brain of summer wakening ground squirrels (WGS), hibernating ground squirrels (HGS), and hibernating ground squirrels with the undercut septum (UHGS) was studied. It was shown that in HGS, the neuropeptides were substantially more effective in modulating the spontaneous activity of MS neurons than in WGS. The undercutting of MS led to the disappearance of the increased responsiveness to the neuropeptides: in UHGS, neuropeptide-induced changes in the spontaneous activity became nearly identical to those in WGS. The decrease in MS responsiveness in UHGS is due mainly to pacemaker neurons, which cease to respond to the peptides. It was shown that the neuropeptides have a dual effect: they change the level of spontaneous activity through direct modulation of pacemaker potential and control responses to electrical stimulation by modulating the synaptic transmission. Contrary to neuropeptides, neurotransmitters were highly effective in neurons of all groups of animals. Presumably, the enhanced excitability of MS during hibernation, which is necessary for performing the 'sentry post' function, is formed under the influence of the preopticohypothalamic area, and this influence is mediated by peptides.     

Postganglionic nerve stimulation induces temporal inhibition of excitability in rabbit sinoatrial node

Vagal stimulation results in complex changes of pacemaker excitability in the sinoatrial node (SAN). To investigate the vagal effects in the rabbit SAN, we used optical mapping, which is the only technology that allows resolving simultaneous changes in the activation pattern and action potentials morphologies. With the use of immunolabeling, we identified the SAN as a neurofilament 160-positive but connexin 43-negative region (n = 5). Normal excitation originated in the SAN center with a cycle length (CL) of 405 +/- 14 ms (n = 14), spread anisotropically along the crista terminalis (CT), and failed to conduct toward the septum. Postganglionic nerve stimulation (PNS, 400-800 ms) reduced CL by 74 +/- 7% transiently and shifted the leading pacemaker inferiorly (78%) or superiorly (22%) from the SAN center by 2-10 mm. In the intercaval region between the SAN center and the septal block zone, PNS produced an 8 +/- 1-mm(2) region of transient hyperpolarization and inexcitability. The first spontaneous or paced excitation following PNS could not enter this region for 500-1,500 ms. Immunolabeling revealed that the PNS-induced inexcitable region is located between the SAN center and the block zone and has a 2.5-fold higher density of choline acetyltransferase than CT but is threefold lower than the SAN center. The fact that the inexcitability region does not coincide with the most innervated area indicates that the properties of the myocytes themselves, as well as intercellular coupling, must play a role in the inexcitability induction. Optically mapping revealed that PNS resulted in transient loss of pacemaker cell excitability and unidirectional entrance conduction block in the periphery of SAN.     

Participation of the genetic apparatus of the hippocampal and neocortical cells in the mechanisms of action of desglycine-arginine vasopressin on learning and memory processes in rats

The paper deals with the influence of synthetic vasopressin analogue--desglycine-arginine vasopressin (DG-AVP) on the content of RNA and fractional composition of chromatin proteins in the tissue of neocortex and hippocampus of intact white rats and after establishing of two-way avoidance reflex. Administration of the peptide alone significantly increased RNA content in hippocampal tissue, injection of the peptide 10 min before conditioning did not lead to significant changes in RNA quantity as compared to that in animals in which the conditioned reflex was established against the background of saline administration. In neocortical tissue neither learning itself nor administration of DG-AVP alone was accompanied by significant changes in RNA content, while learning against the background of peptide injection significantly increased RNA in that structure. In hippocampal and neocortical tissues quantitative changes were observed in certain fractions of chromatin proteins in all animal groups studied.     

Bioelectric activity of various parts of the upper urinary tract

Experiments on 50 dogs carried out by the method of in vivo electromyography have confirmed the localization of the pacemaker of the pyeloureter in the proximal end of the renal pelvis at the pelvicalyceal border. It is determined that the pacemaker biopotentials are characterized by higher frequency, low-voltage rhythmic amplitude, special form and the highest stability of parameters to the diuresis changes and adrenalin influence. Biopotentials are spreading in the distal direction with an increasing speed, amplitude and decreasing frequency. Pyeloureteral junction is the main rhythmoregulator of the bioelectric activity of the ureter. Synchronization of the bioelectric activity of different parts of the pyeloureter depends on the level of its excitability.     

NALCN: A Regulator of Pacemaker Activity

Pacemaker cells play a fundamental role in generating or regulating many essential biological rhythms. Spontaneous pacemaker activity is dependent on the function of an array of ion channels expressed in these cells. Recent characterization of a Na(+) leak channel (NALCN) has linked to its role in conducting the background Na(+) current that depolarizes resting membrane properties of pacemaker neurons. NALCN, along with Unc79 and Unc80, forms a protein complex that is involved in regulating intrinsic membrane and synaptic activities. In this review, we will discuss the current understanding of NALCN channel physiology and its role in regulating cell excitability and pacemaker activity.     

Ether-a-go-go-related gene 3 is the main candidate for the E-4031-sensitive potassium current in the pacemaker interstitial cells of Cajal

The interstitial cells of Cajal (ICC), as pacemaker cells of the gut, contribute to rhythmic peristalsis and muscle excitability through initiation of slow-wave activity, which subsequently actively propagates into the musculature. An E-4031-sensitive K(+) current makes a critical contribution to membrane potential in ICC. This study provides novel features of this current in ICC in physiological solutions and seeks to identify the channel isoform. In situ hybridization and Kit immunohistochemistry were combined to assess ether-a-go-go-related gene (ERG) mRNA expression in ICC in mouse jejunum, while the translated message was assessed by immunofluorescence colocalization of ERG and Kit proteins. E-4031-sensitive currents in cultured ICC were studied by the whole cell patch-clamp method, with physiological K(+) concentration in the bath and the pipette. In situ hybridization combined with Kit immunohistochemistry detected m-erg1 and m-erg3, but not m-erg2, mRNA in ICC. ERG3 protein was colocalized with Kit-immunoreactive ICC in jejunum sections, but ERG1 protein was visualized only in the smooth muscle cells. At physiological K(+) concentration, the E-4031-sensitive outward current in ICC was different from its counterpart in cardiac and gut smooth muscle cells. In particular, inactivation upon depolarization and recovery from inactivation by hyperpolarization were modest in ICC. In summary, the E-4031-sensitive currents influence the kinetics of the pacemaker activity in ICC and contribute to maintenance of the resting membrane potential in smooth muscle cells, which together constitute a marked effect on tissue excitability. Whereas this current is mediated by ERG1 in smooth muscle, it is primarily mediated by ERG3 in ICC.     

Effect of chronic disconnection between septal area and hypothalamus on modulation of background activity of septal neurons by biologically-active substances in hibernators

Our previous work demonstrated paradoxically increased excitability of the medial septal (MS) neurons during hibernation of ground squirrels in comparison to waking animals. Recently this was supported by demonstration of higher efficacy of the neuropeptides identified in the brain of hibernators in septal slices of hibernating animals. To decide whether this increased excitability is determined by endogenous properties of the pacemaker septal neurons, or it depends on the influences of thermoregulatory-circadian mechanisms of preoptico-hypothalamic area, testing of the neuropeptides (TSKYR, TSKY, DY) and neurotransmitters participating in control of hibernation (serotonin and noradrenaline) was repeated on septal slices taken from the brain of hibernating animals two weeks after operation disconnecting it from the hypothalamus. Effects of neuropeptides in the deafferented hibernating animals neither quantitatively (low reactivity level), nor qualitatively (distribution of inhibitory and excitatory responses) differed from the data obtained in waking animals. Decrease of reactivity occurred at the expense of the neurons with regular pacemaker-like spontaneous activity. Thus, increased reactivity of the MS neurons to neuropeptides in hibernating animals depends mainly on influence of the hypothalamic centres controlling hibernation behavior upon pacemaker neurons of the MS. Contrary to the neuropeptides, serotonin and noradrenaline were highly effective in deafferented septum. They evoked stronger changes of background activity (shorter latencies and more rapid development of maximal shifts), presumably as a result of development of denervation hypersensitivity after deafferentation.     

ERG K+ currents regulate pacemaker activity in ICC

Ether-à-go-go-related gene (ERG) K channels have been implicated in the generation of pacemaker activities in the heart. To study the presence and function of ERG K channels in the pacemaker cells of the small intestine [the interstitial cells of Cajal (ICC)], a combination of patch-clamp techniques, tissue and live cell immunohistochemistry, RT-PCR, and in vitro functional studies were performed. Nonenzymatically isolated ICC in culture were identified by vital staining and presence of rhythmic inward currents. RT-PCR showed the presence of ERG mRNA in the intestinal musculature, and immunohistochemistry on tissue and cultured cells demonstrated that protein similar to human ERG was concentrated on ICC in the Auerbach's plexus region. Whole cell ERG K+ currents were evoked on hyperpolarization from 0 mV (but not from -70 mV) up to -120 mV and showed strong inward rectification. The currents were inhibited by E-4031, cisapride, La3+, and Gd3+ but not by 50 microM Ba2+. The ERG K+ inward current had a typical transient component with fast activation and inactivation kinetics followed by significant steady-state current. E-4031 also inhibited tetraethylammonium (TEA)-insensitive outward current indicating that the ERG K+ current is operating at depolarizing potentials. In contrast to TEA, blockers of the ERG K+ currents caused marked increase in tissue excitability as reflected by an increase in slow-wave duration and an increase in superimposed action potential activity. In summary, ERG K channels in ICC contribute to the membrane potential and play a role in regulation of pacemaker activity of the small intestine.     

Action of pilocarpine on the normal frog heart and in pathology

Experiments on frogs were performed to examine the effect of the M-cholinomimetic pilocarpine on the heart. It was discovered that at concentrations of 10(-15)--10(-5) g/ml pilocarpine exerted only an adverse chronotropic effect on the perfused heart. When applied at a concentration of 10(-4) g/ml the drug produced a negative as well as a positive chronotropic effect. The latter occurred spasmodically (without progressive rise in the heart rate) in association with a slow heart rate. In some experiments such effects were preceded by a certain deceleration of the heart. In experiments with positive chronotropic effects, arrhythmias and sinoatrial dissociation were observed sometimes. Experiments with recording of the electrograms of the sinuses and lower parts showed that such effects were caused not by pacemaker acceleration but by the removal of the blockade of conduction, between the pacemaker and the atria. As far as the pacemaker is concerned, pilocarpine exerted only a negative chronotropic effect.     

Temperature acclimation modifies sinoatrial pacemaker mechanism of the rainbow trout heart

The hypothesis of pacemaker level origin of thermal compensation in heart rate was tested by recording action potentials (AP) in intact sinoatrial tissue and enzymatically isolated pacemaker cells of rainbow trout acclimated at 4 degrees C (cold) and 18 degrees C (warm). With electrophysiological recordings, the primary pacemaker was located at the base of the sinoatrial valve, where a morphologically distinct ring of tissue comprising myocytes and neural elements was found by histological examination. Intrinsic beating rate of this pacemaker was higher in cold-acclimated (46 +/- 6 APs/min) than warm-acclimated trout (38 +/- 3 APs/min; P < 0.05), and a similar difference was seen in beating rate of isolated pacemaker cells (44 +/- 6 vs. 38 +/- 6 APs/min; P < 0.05), supporting the hypothesis that thermal acclimation modifies the intrinsic pacemaker mechanism of fish heart. Inhibition of sarcoplasmic reticulum (SR) with 10 microM ryanodine and 1 microM thapsigargin did not affect heart rate in either warm- or cold-acclimated trout at 11 degrees C but reduced heart rate in warm-acclimated trout from 74 +/- 2 to 42 +/- 6 APs/min (P < 0.05) at 18 degrees C. At 11 degrees C, a half-maximal blockade of the delayed rectifier K+ current (I(Kr)) with 0.1 microM E-4031 reduced heart rate more in warm-acclimated (from 45 +/- 1 to 24 +/- 5 APs/min) than cold-acclimated trout (56 +/- 3 vs. 48 +/- 2 APs/min), whereas I(Kr) density was higher and AP duration less in cold-acclimated trout (P > 0.05). Collectively, these findings suggest that a cold-induced increase in AP discharge frequency is at least partly due to higher density of the I(Kr) in the cold-acclimated trout, whereas contribution of SR Ca2+ release to thermal compensation of heart rate is negligible.