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Biomedical subjects

G D Hirst

Publications and source records attributed to G D Hirst.

At least 19 recordsLinked to original sources
PubMed2001

Failure of Ba2+ and Cs+ to block the effects of vagal nerve stimulation in sinoatrial node cells of the guinea-pig heart.

The aim of the study was to evaluate which ionic currents are modified in the sinoatrial node of guinea pigs when the vagus is stimulated. Responses of isolated atrial preparations to bilateral vagus nerve stimulation were examined. In bath-mounted preparations, 10-s trains of vagal stimulation (1-50 Hz) slowed the rate at which atrial contractions occurred. After the trains of stimuli, the force generated by each contraction was reduced. Both vago-inhibitory responses persisted in the presence of caesium (2 mM) and barium ions (1 mM). Vagal stimulation evoked a similar bradycardia in superperfused preparations in which intracellular recordings were made from pacemaker cells in the sinoatrial node. When pacemaking activity was abolished by adding the organic calcium channel antagonist nifedipine (1 microM) to the perfusate, vagal stimulation generated an inhibitory junction potential (IJP). Both the bradycardia and the amplitude of the inhibitory junction potential increased as the frequency of vagal stimulation was increased. The ability of vagal stimulation to produce inhibitory junction potentials was unaffected by the addition of caesium and barium ions to the perfusate. These observations suggest that the negative chronotropic and inotropic responses to vagal stimulation only minimally involve a muscarinically activated potassium current (I(KACh)) or changes in the hyperpolarization-activated pacemaker current Ih.

Action Potentials↗
PubMed2001

Generation of slow waves in the antral region of guinea-pig stomach--a stochastic process.

1. Slow waves were recorded from the circular muscle layer of the antral region of guinea-pig stomach. Slow waves were abolished by 2APB, an inhibitor of IP(3)-induced Ca2+ release. 2. When the rate of generation of slow waves was monitored it was found to vary from cycle to cycle around a mean value. The variation persisted after abolishing neuronal activity with tetrodotoxin. 3. When simultaneous recordings were made from interstitial cells in the myenteric region (ICC(MY)) and smooth muscle cells of the circular layer, variations in the rate of generation of slow waves were found to be linked with variations in the rate of generation of driving potentials by ICC(MY). 4. A preparation was devised which consisted of the longitudinal muscle layer and ICC(MY). In this preparation ICC(MY) and smooth muscle cells lying in the longitudinal muscle layer generated driving potentials and follower potentials, synchronously. 5. Driving potentials had two components, a rapid primary component that was followed by a prolonged plateau component. Caffeine (3 mM) abolished the plateau component; conversely reducing the external concentration of calcium ions [Ca2+](o) mainly affected the primary component. 6. Analysis of the variations in the rate of generation of driving potentials indicated that this arose because both the duration of individual driving potentials and the interval between successive driving potentials varied. 7. It is suggested that the initiation of pacemaker activity in a network of ICC(MY) is a stochastic process, with the probability of initiating a driving potential slowly increasing, after a delay, from a low to a higher value following the previous driving potential.

Animals↗
PubMed2001

Selective knockout of intramuscular interstitial cells reveals their role in the generation of slow waves in mouse stomach.

1. Intracellular recording techniques were used to compare the patterns of electrical activity generated in the antral region of the stomachs of wild-type and W/W(V) mutant mice. Immunohistochemical techniques were used to determine the distribution of c-kit-positive interstitial cells of Cajal (ICC) within the same region of the stomach. 2. In wild-type mice interstitial cells were found at the level of the myenteric plexus (ICC(MY)) and distributed within the smooth muscle bundles (ICC(IM)). In these preparations slow waves, which consisted of initial and secondary components, were detected. 3. In W/WV mutant mice ICC(MY) could be identified at the level of the myenteric plexus but ICC(IM) were not detected within smooth muscle bundles. Intracellular recordings revealed that smooth muscle cells generated waves of depolarization; these lacked a secondary component. 4. These results indicate that the secondary regenerative component of a slow wave is generated by ICC(IM). Thus the depolarization arising from the pacemaker cells, ICC(MY), is augmented by ICC(IM), so causing a substantial membrane depolarization in the circular muscle layer. Rather than contributing directly to rhythmical electrical activity, smooth muscle cells appear to depolarize at the command of the two subpopulations of ICC.

Animals↗
PubMed2001

Parallel metabotropic pathways in the heart of the toad, Bufo marinus.

This study examined the transduction pathways activated by epinephrine in the pacemaker region of the toad heart. Recordings of membrane potential, force, and intracellular Ca(2+) concentration ([Ca(2+)](i)) were made from arrested toad sinus venosus. Sympathetic nerve stimulation activated non-alpha-, non-beta-adrenoceptors to evoke a membrane depolarization and a transient increase in [Ca(2+)](i). In contrast, the beta-adrenoceptor agonist isoprenaline (10 microM) caused membrane hyperpolarization and decreased [Ca(2+)](i). The phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (0.5 mM) mimicked the isoprenaline-evoked membrane hyperpolarization. Epinephrine (10-50 microM) caused an initial membrane depolarization and an increase in [Ca(2+)](i) followed by membrane hyperpolarization and decreased [Ca(2+)](i). The membrane depolarizations evoked by sympathetic nerve stimulation or epinephrine were abolished either by the phospholipase C inhibitor U-73122 (20 microM) or by the blocker of D-myo-inositol 1,4,5,-trisphosphate-induced Ca(2+) release, 2-aminoethoxydiphenyl borate (2-APB, 60 microM). Neither U-73122 nor 2-APB had an affect on the membrane hyperpolarization evoked by beta-adrenoceptor activation. These results suggest that in the toad sinus venosus, two distinct transduction pathways can be activated by epinephrine to cause an increase in heart rate.

Adrenergic Agonists↗
PubMed2000

Potentiation by neostigmine of responses to vagal nerve stimulation in the sinus venosus of the toad.

The effects of the cholinesterase inhibitor neostigmine on the responses to vagus nerve stimulation of isolated sinus venosus/atrial preparations of the toad, Bufo marinus, were examined. In control solutions, trains of stimuli applied to the vagus nerve led to a decrease in heart rate that was susceptible to muscarinic receptor blockade. Membrane potential recordings made from sinus venosus cells showed that the responses to trains of stimuli, delivered at frequencies of less than 10 Hz, were little changed by the addition of neostigmine. However, the responses to longer trains of stimuli at 10 Hz (30 versus 10 s) were potentiated and the nature of the membrane potential changes was altered. The results suggest that, due to the activity of cholinesterases, acetylcholine (ACh) released from parasympathetic nerves normally has little access to the muscarinic receptors in the pacemaker region, which are linked to potassium channels.

Acetylcholine↗
PubMed2000

Mechanisms of excitatory neuromuscular transmission in the guinea-pig urinary bladder.

1. In smooth muscle of the guinea-pig bladder, either membrane potential recordings or [Ca2+]i measurements were made simultaneously with isometric tension recordings. 2. Single transmural stimuli initiated excitatory junction potentials (EJPs) which triggered action potentials, transient increases in [Ca2+]i and associated contractions. These responses were abolished by alpha, beta-methylene ATP, suggesting that they resulted from the activation of purinoceptors by neurally released ATP. 3. Nifedipine abolished action potentials leaving the underlying EJPs and reduced the amplitude of both nerve-evoked increases in [Ca2+]i and associated contractions. The subsequent co-application of caffeine and ryanodine inhibited the residual responses without inhibiting EJPs. These results indicate that stimulation of purinoceptors activates both Ca2+ influx through L-type Ca2+ channels and Ca2+ release from intracellular Ca2+ stores. 4. In the presence of alpha, beta-methylene ATP, trains of stimuli failed to initiate EJPs but increased the frequency of action potentials. Trains of stimuli also initiated oscillatory increases in [Ca2+]i and associated contractions. These responses were abolished by hyoscine, indicating that they resulted from the activation of muscarinic receptors by neurally released ACh. 5. Oscillatory increases in [Ca2+]i and associated contractions were inhibited by either nifedipine or caffeine, indicating that the stimulation of muscarinic receptors activates both Ca2+ influx through L-type Ca2+ channels and Ca2+ release from intracellular Ca2+ stores.

Action Potentials↗
PubMed2000

Vagal inhibition in the antral region of guinea pig stomach.

The effects of vagal stimulation in the presence of a muscarinic antagonist were examined on three distinct rhythmically active cells located in guinea pig antrum. Vagal stimulation inhibited contractions of the circular muscle layer but did not change their rate of occurrence. With the use of intracellular recording techniques, these stimuli were found to initiate inhibitory junction potentials in the circular layer but produced smaller potential changes in driving and follower cells. Inhibition of the circular muscle layer involved two separate components. The dominant component was independent of changes in membrane potential and was abolished by nitro-L-arginine. After abolishing Ca(2+) entry into smooth muscle cells with a Ca(2+) antagonist, vagal stimulation continued to inhibit the residual contractions associated with each slow wave. When the cyclic changes in intracellular Ca(2+) concentration associated with each slow wave were measured, they were found to be unchanged by vagal stimulation. The observations suggest that vagal inhibition of stomach movements does not alter pacemaker activity in the stomach; rather, it results from a change in the sensitivity of smooth muscle contractile proteins to Ca(2+).

Animals↗
PubMed1999

Unitary nature of regenerative potentials recorded from circular smooth muscle of guinea-pig antrum.

1. When short segments of single bundles of circular muscle of guinea-pig antrum were isolated and impaled with two microelectrodes, the membrane potential recordings displayed an ongoing discharge of noise. 2. Treating the preparations with acetoxymethyl ester form of BAPTA (BAPTA AM) reduced the membrane noise and revealed discrete depolarizing unitary potentials. The spectral densities determined from control preparations and ones loaded with BAPTA had similar shapes but those from control preparations had higher amplitudes, suggesting that membrane noise results from a high frequency discharge of unitary potentials. 3. Depolarization of isolated segments of antrum initiated regenerative responses. These responses, along with membrane noise and unitary potentials, were inhibited by a low concentration of caffeine (1 mM). 4. Loading the preparations with BAPTA decreased the amplitudes of regenerative responses. Depolarization was now seen to increase the frequency and mean amplitude of unitary potentials over a time course similar to that of a regenerative potential. 5. Noise spectra determined during periods of rest, during regenerative potentials triggered by direct depolarization and during slow waves, recorded from preparations containing interstitial cells of Cajal (ICC), had very similar shapes but different amplitudes. 6. The observations suggest that a regenerative potential, the secondary component of a slow wave, is made up of a cluster of several discrete unitary potentials rather than from the activation of voltage-dependent ion channels.

Animals↗
PubMed1999

Regenerative potentials evoked in circular smooth muscle of the antral region of guinea-pig stomach.

1. Slow waves recorded from the circular smooth muscle layer of guinea-pig antrum consisted of two components, an initial component and a secondary regenerative component. Whereas both components persisted in the presence of nifedipine, the secondary component was abolished by a low concentration of caffeine. 2. Short segments of single bundles of circular muscle were isolated and impaled with two microelectrodes. Depolarizing currents initiated regenerative responses which resembled those initiated during normal slow waves. These responses had partial refractory periods of 20-30 s and were initiated about 1 s after the onset of membrane depolarization. 3. The regenerative responses persisted in the presence of either nifedipine or cobalt ions but were abolished by caffeine, BAPTA or cyclopiazonic acid. 4. The observations suggest that depolarizing membrane potential changes trigger the release of Ca2+ from intracellular stores and this causes a depolarization by activating sets of unidentified ion channels in the membranes of smooth muscle cells of the circular layer of guinea-pig antrum.

Animals↗
PubMed1999

Sympathetic neuroeffector transmission in the rat anococcygeus muscle.

1. When intracellular recordings were made from preparations of rat anococcygeus muscle, transmural nerve stimulation evoked noradrenergic excitatory junction potentials (EJPs) made up of two distinct components. Both components were abolished by either guanethidine or alpha-adrenoceptor antagonists, indicating that they resulted from the release of transmitter from sympathetic nerves and the subsequent activation of alpha-adrenoceptors. 2. The first component was associated with a transient increase in the intracellular concentration of calcium ions ([Ca2+]i) and a contraction. Although the second component was often associated with a long lasting increase in [Ca2+]i it was not associated with a contraction unless the second component initiated an action potential. 3. The increase in [Ca2+]i associated with the first component resulted from Ca2+ release from an intracellular store and from entry of Ca2+ through voltage-dependent Ca2+ channels. The increase in [Ca2+]i associated with the second component resulted only from the entry of Ca2+ through L-type Ca2+ channels (CaL channels). The depolarization associated with the initial increase in [Ca2+]i was abolished by reducing the external concentration of chloride ions ([Cl-]o), suggesting that it involved the activation of a Cl- conductance. 4. When the relationships between changes in [Ca2+]i, membrane depolarization and contraction produced by an increasing number of sympathetic nerve stimuli were determined in control, and caffeine- and nifedipine-containing solutions, it was found that an increase in [Ca2+]i recorded in nifedipine produced a larger contraction and larger membrane depolarization than did a similar increase in [Ca2+]i recorded in either control or caffeine-containing solutions. These observations indicate that Ca2+ released from stores more readily triggers contraction and membrane depolarization than does Ca2+ entry via CaL channels.

Action Potentials↗
PubMed1999

Identification of rhythmically active cells in guinea-pig stomach.

1. When intracellular recordings were made from the antral region of guinea-pig stomach, cells with different patterns of electrical activity were detected. 2. One group of cells, slow-wave cells, generated slow waves which consisted of initial and secondary components. When filled with either Lucifer Yellow or neurobiotin, the cells identified as smooth muscle cells lying in the circular muscle layer. 3. A second group of cells, driving cells, generated large, rapidly rising, potential changes, driving potentials. They had small cell bodies with several processes. With neurobiotin, a network of cells was visualized that resembled c-kit positive interstitial cells of the myenteric region. 4. A third group of cells generated sequences of potential changes which resembled driving potentials but had smaller amplitudes and slow rates of rise. These cells resembled smooth muscle cells lying in the longitudinal muscle layer. 5. When simultaneous recordings were made from the driving and slow-wave cells, driving potentials and slow waves occurred synchronously. Current injections indicated that both cell types were part of a common electrical syncytium. 6. The initial component of slow waves persisted in low concentrations of caffeine, but the secondary component was abolished; higher concentrations shortened the duration of the residual initial component. Driving potentials continued in the presence of low concentrations of caffeine; moderate concentrations of caffeine shortened their duration. 7. Hence three different types of cells were distinguished on the basis of their electrical activity, their responses to caffeine and their structure. These were smooth muscle cells, lying in the longitudinal and circular layers, and interstitial cells in the myenteric region. The observations suggest that interstitial cells initiate slow waves.

Animals↗
PubMed1998

Regional differences in sympathetic neurotransmission to cutaneous arteries in the guinea-pig isolated ear.

The effects of sympathetic nerve stimulation on different cutaneous arteries were examined in arteries isolated from guinea-pig ears, by measuring membrane potential changes in smooth muscle cells in response to electrical field stimulation. Resting membrane potential (RMP) was similar in proximal (main ear artery) and distal (3rd or 4th branch order) cutaneous arteries (mean -71 mV). Single stimuli evoked excitatory junction potentials (EJPs) in all arteries. The EJPs in proximal arteries were twice the amplitude, and the time constant of EJP decay was almost half the value, compared with distal cutaneous arteries. EJP amplitude was reduced by > 90% by suramin (30 microM) or alpha,beta,methylene-ATP (alpha,beta,m-ATP)(1 microM) in all proximal, and most distal arteries. Residual responses in distal arteries were resistant to tetrodotoxin. The N-type calcium channel blocker, omega-conotoxin GVIA (30 nM), reduced EJP amplitude by 70-100% in both proximal and distal arteries. Successive EJPs evoked by trains of stimuli at 1 to 5 Hz were depressed in amplitude in proximal arteries, but showed facilitation in distal arteries. EJP depression in proximal arteries was reversed to facilitation by the alpha2-adrenoceptor antagonist, yohimbine (30 nM). Trains of stimuli delivered at 10-20 Hz produced summation of EJPs and active membrane responses in 30% of proximal arteries. Active responses were never detected in distal arteries. Slow depolarizations following the EJPs were detected in most arteries after trains of stimuli, and were abolished by prazosin (0.3 microM) or omega-conotoxin GVIA (30 nM). The density of the perivascular plexus of axons innervating proximal arteries, demonstrated with catecholamine fluorescence histochemistry, was twice that in distal cutaneous arteries. These regional differences in sympathetic neurotransmission suggest that cutaneous vasoconstriction in response to thermoregulatory stimuli, which occurs predominantly in distal cutaneous segments, is likely to be qualitatively different from cutaneous vasoconstriction of proximal arteries in response to other physiological stimuli.

Adrenergic alpha-Antagonists↗
PubMed1997

Distribution of synaptic boutons around identified neurones lying in the cardiac plexus of the guinea-pig.

Neurones in the cardiac plexus of the guinea-pig were subdivided into three groups according to their electrophysiological properties and subsequently labelled with neurobiotin. Preparations were counterstained with antibodies to synaptophysin to reveal boutons containing synaptic vesicles. Two of the three groups of cells which, in the electrophysiological studies were found to receive excitatory synaptic inputs were associated with numerous synaptophysin positive boutons. Cells of the other group, which did not appear to receive any synaptic inputs were associated with fewer synaptophysin positive boutons. Wholemount preparations were double stained with antibodies to Protein Gene Product 9.5 and synaptophysin. This revealed most neurones, along with vesicle filled boutons. Ganglia generally contained less than 10 neurones with a range between one and over 30. In wholemount preparations it was found that about 90% of the cells were associated with a large number of synaptophysin positive boutons whilst the remaining cells were associated with very few synaptophysin positive boutons. These results are consistent with the idea that a proportion of cardiac ganglion cells fail to receive a synaptic input and suggest that by staining wholemount preparations of guinea-pig cardiac plexus with antibodies to synaptophysin it is possible to differentiate between groups of cells which receive a synaptic input and those which fail to receive a synaptic input. Approximately 10% of intrinsic cardiac neurones appear to lack a synaptic input.

Animals↗
PubMed1997

Electrical properties of iridial arterioles of the rat.

When intracellular recordings were made from iridial arterioles, the cells had membrane potentials of about -65 mV and perivascular nerve stimulation evoked a membrane depolarization. When these cells were labeled with lucifer yellow, all cells that responded to perivascular nerve stimulation had the morphological characteristics of smooth muscle cells. Cells with the morphological characteristics of endothelial cells were never stained. When impaled with two separate recording electrodes, the smooth muscle layer was shown to form an electrical syncytium with a membrane time constant of approximately 80 ms and an electrical length constant of approximately 900 microns. At the ultrastructural level, areas of close apposition were frequently observed between adjacent smooth muscle cells and between adjacent endothelial cells. On the other hand, at contacts between smooth muscle and endothelial cells, the membranes characteristically had much larger separations. The observations show that individual smooth muscle cells are electrically coupled to their neighbors, but the morphological studies raise the possibility that in these arterioles the endothelial and muscle layers are electrically separate.

Animals↗
PubMed1996

Differences in the responses to purinergic nerve stimulation and applied ATP in the guinea-pig vas deferens.

The responses to sympathetic nerve stimulation and to applied ATP in the guinea-pig vas deferens were compared. Nifedipine (10 microM) markedly reduced the non-adrenergic neural contraction but only partially blocked the contractions produced by bath-applied ATP. Suramin (300 microM) also markedly reduced the contractile responses produced by nerve stimulation, but had no significant effect on the contractions produced by bath-applied ATP. Using intracellular recording techniques, nerve stimulation was shown to produce an excitatory junction potential which was abolished by suramin (1 microM). Ionophoretic application of ATP and bath-applied ATP also produced a depolarization. Suramin (1 microM) failed to abolish the response to bath-applied ATP and enhanced the ionophoretically induced depolarization. These results suggest either that ATP is not a transmitter in the vas deferens or that two classes of purinoceptor are present, one suramin-sensitive receptor which produces a contraction via the opening of voltage-dependent Ca2+ channels, and another which is suramin-resistant and produces a contraction by another means.

Adenosine Triphosphate↗