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Biomedical subjects

A F Brading

Publications and source records attributed to A F Brading.

At least 109 records · Page 6Linked to original sources
PubMed1989

The effects of cromakalim on the detrusor muscle of human and pig urinary bladder.

The effects of cromakalim, a potassium channel activating drug, have been studied on isolated detrusor muscle strips from normal Landrace boar, normal and unstable mini-pig and unstable human bladder. Cromakalim abolished spontaneous activity in all strips but did not abolish the ability of the detrusor muscle from any of the specimens studied to respond to carbachol, increased extracellular K+ or transmural nerve stimulation. Intravenous infusion of cromakalim in the urethral obstructed mini-pig caused the characteristic unstable contractions associated with bladder outflow obstruction to be abolished, leaving the animal able to void. Experiments with potassium isotopes and the sucrose gap technique demonstrated that cromakalim increased the potassium permeability and hyperpolarised the cell membrane, consistent with its reported actions on other smooth muscles. These results suggest that drugs such as cromakalim, which act by reducing membrane excitability without inhibiting responses to existing innervation, may have a clinical application in the treatment of instability which is secondary to bladder outflow obstruction.

Animals↗
PubMed1989

Sequential morphological changes in the pig detrusor in response to chronic partial urethral obstruction.

The sequential morphological changes which occur in the wall of the porcine urinary bladder in response to experimentally induced partial outflow obstruction have been determined using morphometric light and electron microscope techniques. The initial morphological response to urethral obstruction was a reduction of approximately 50% in the density of autonomic nerves in the bladder wall which occurred within the first 3 months. During this period the morphology of smooth muscle cells and the distribution of connective tissue were unaltered. Longer periods of obstruction produced a gradual further reduction in the density of innervation. However, from 3 months onwards the detrusor muscle bundles became infiltrated by connective tissue, although significant change in muscle cell size was not detected after obstruction for 12 months. Using electron microscopy, smooth muscle cells were observed to possess extensive amounts of perinuclear granular reticulum, this feature being particularly marked in bladders subjected to longer periods of obstruction. The underlying mechanisms responsible for these structural changes remain to be determined.

Animals↗
PubMed1989

The effect of cromakalim on the smooth muscle of the guinea-pig urinary bladder.

1. The actions of cromakalim were studied on the detrusor muscle from guinea-pig urinary bladder. Cromakalim reduced the frequency and amplitude of spontaneous contractile activity of the smooth muscle of the guinea-pig urinary bladder at 5 x 10(-8)M and abolished the activity at concentrations above 5 x 10(-7)M. 2. Electrophysiological experiments demonstrated that cromakalim increased membrane conductance, caused a dose-dependent hyperpolarization of the cell membrane and loss of spike activity. These events are consistent with the opening of K+ channels. 3. The effects of 10(-6)M and 10(-5)M cromakalim on the contractile responses to carbachol, potassium and transmural nerve stimulation were studied. Cromakalim did not prevent the detrusor from responding to these agents, although it significantly reduced the contractile response to K+ at concentrations below 70 mM. 4. Uptake and efflux experiments using 86Rb+ were unable to demonstrate any significant effect on transmembrane movement produced by cromakalim (10(-5)M). 5. 43K+ efflux showed a dose-dependent increase in the rate constant on addition of cromakalim. The difference in the selectivity for K+ over Rb+ was confirmed in dual label uptake experiments. 6. Substitution experiments in which the K+ ions in the tissue were gradually replaced by Rb+ demonstrated that cromakalim had a progressively decreasing effect on spontaneous activity as internal K+ was lowered. When all the K+ was replaced by Rb+, cromakalim no longer inhibited spontaneous activity, confirming that the channel opened by cromakalim appears relatively impermeant to Rb+.

Animals↗
PubMed1989

Electrical and mechanical responses of guinea-pig bladder muscle to nerve stimulation.

1 The electrical and mechanical responses to transmural stimulation of intrinsic nerves have been recorded from smooth muscle strips dissected from the dome of the guinea-pig bladder, by use of intracellular microelectrodes, and conventional tension recording techniques. 2 Stimulation of intrinsic nerves evoked action potentials in all cells studied. Hyperpolarization of the cells by extracellular current injection revealed subthreshold excitatory junction potentials (e.j.ps) in about a quarter of the cells studied. 3 Action potentials could still be evoked in the presence of atropine and neostigmine, but were abolished after desensitization of the cells to alpha, beta-methylene ATP, a stable analogue of ATP. 4 In the presence of neostigmine, the evoked action potential was followed by a slow depolarization of the membrane. The mechanical response increased in amplitude and duration. 5 The contractile response to transmural nerve stimulation was reduced but not abolished in the presence of either atropine or desensitizing doses of alpha, beta-methylene ATP. Atropine was more effective at high frequencies of stimulation (greater than or equal to 30 Hz), and alpha, beta-methylene ATP at low frequencies (less than or equal to 15 Hz). In combination the drugs abolished the response. 6 The results suggest that the mechanical response to excitatory nerve stimulation is biphasic. The early transient response is elicited by e.j.ps and evoked spikes, is resistant to atropine, but sensitive to desensitization of purinoceptors. The late response is mediated through muscarinic receptors, involves little membrane depolarization, and is unaffected by desensitization of purinoceptors. These responses are analogous to the responses seen in rabbit bladder, and in the sympathetically innervated rat tail artery and guinea-pig vas deferens.

Acetylcholine↗
PubMed1988

An in vitro pharmacological study of the human trigone--a site of non-adrenergic, non-cholinergic neurotransmission.

Muscle strips from the human detrusor and trigone were studied in vitro. The detrusor muscle contracted strongly to both cholinergic receptor stimulation with carbachol and to electrical field stimulation. There was no evidence of atropine resistance in the detrusor strips. The superficial trigone responded maximally to alpha-adrenergic receptor stimulation but also produced a significant cholinergic response. Intramural nerve stimulation in the presence of both atropine and phentolamine produced a residual non-adrenergic, non-cholinergic (NANC) response of 40% of its maximum at 5 Hz. Electrical stimulation, particularly at the lower frequencies of stimulation, produced relaxation responses in 40% of the superficial trigonal muscle strips. These relaxations were not blocked by atropine, phentolamine or propranolol, but were abolished by tetrodotoxin. The possible role of the cholinergic "input" to the superficial trigone and the importance of the NANC excitatory and inhibitory innervation in preventing vesico-ureteric reflux and and in aiding bladder neck opening is discussed.

Atropine↗
PubMed1988

Contractile effects of manganese on taenia of guinea pig cecum.

The present study was undertaken to examine Mn as a contractile agent in its own right in the absence of extracellular Ca and after the depletion of Ca from agonist-releasable stores. The results show that isolated strips of taenia coli depolarized by high-K, Ca-free solution will contract when Mn is present. Similarly, carbachol will evoke contractions in tissues incubated in Ca-free, Mn-containing solutions. In addition, it seems possible to stimulate contraction by releasing Mn from an agonist-releasable intracellular store in the absence of extracellular Ca and Mn. Therefore the possibility exists that cations employed as Ca antagonists may in fact support contraction.

Animals↗
PubMed1987

Bladder outflow obstruction--a cause of denervation supersensitivity.

Eighteen Landrace pigs and 12 Göttingen mini-pigs were evaluated in a study of experimental bladder outflow obstruction. Twenty-two of the animals underwent partial bladder outflow obstruction for periods up to 12 months. The subsequent changes were assessed using cystometric, physiological and morphological means. There was a consistent increase in the voiding pressures and a concomitant reduction in the flow rates in all the obstructed animals. Seventy-seven per cent of the obstructed animals showed cystometric evidence of bladder instability. In vitro studies showed an increase in sensitivity to exogenously applied agonists and a reduction in sensitivity to intramural nerve stimulation. Morphological studies showed an inverse correlation between neuronal density and the duration of obstruction. These changes are typical of post-junctional supersensitivity secondary to partial denervation. These results suggest that agents capable of stabilising the bladder smooth muscle membrane may be useful in the treatment of detrusor instability secondary to bladder outflow obstruction.

Animals↗
PubMed1987

An investigation of sodium-calcium exchange in the smooth muscle of guinea-pig ureter.

1. After application of ouabain (10(-4) M), the intracellular Na+ activity (alpha iNa) of smooth muscle cells in the guinea-pig ureter stabilizes at a relatively low level which can be rapidly lowered by reduction of external Na+ (Na+o) or elevation of Ca2+o. Both these procedures also elicit a transient contracture. These observations have previously been interpreted as evidence for Na+-Ca2+ exchange. The presence of such an exchange mechanism has now been further investigated by measurements of alpha iNa, tension, ion analysis and 22Na efflux. 2. Ion analysis demonstrated that tissues were able to maintain a high cellular K+ content in the presence of ouabain, but slowly lost K+ and gained Na+ if K+o was also removed, as expected for an infinite outward gradient for K+ and a fully inhibited Na+ pump. 3. Tissues were only able to maintain a low cellular Na+ and high cellular K+ in the presence of ouabain if Ca2+ was present in the bathing solution. Reduction of Ca2+o to very low levels also caused a continual slow rise in alpha iNa in the presence of ouabain, provided that the prolonged depolarization caused by these low levels was prevented by elevation of Mg2+o. Alteration of the membrane potential by changing K+o at constant Na+o showed that alpha iNa decreased by about 1.2 mM for a 10 mV depolarization, within the range from -70 to -30 mV. 4. A small Ca2+o-activated 22Na efflux was observed in ouabain-treated tissues in the absence of Na+o. 40 mM-Ca2+ was not more effective at activating this efflux than was 2.5 mM-Ca2+, while 40 mM-Mg2+ was ineffective. Restoration of the normal Na+o caused a large increase in the rate of 22Na loss. 5. Application of Mn2+ in the presence of ouabain caused a slow rise in alpha iNa and a small decline in resting tension. The fall in alpha iNa on reduction of Na+o was slowed by the presence of Mn2+ (mean half-time increased from 1.7 to 5.0 min) and the concomitant contracture was almost abolished. These results are consistent with a Mn2+-induced inhibition of Na+-Ca2+ exchange. However, the fall in alpha iNa induced by elevation of Ca2+o was unaffected by the presence of Mn2+ and the attendant contracture was, if anything, enhanced. 6. Observation of changes in alpha iNa and tension at various Mn2+ and Ca2+ concentrations demonstrated a competitive interaction between the two divalent cations.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗
PubMed1985

Na-Ca exchange in vascular smooth muscle.

The above section makes it clear that even if an Na-Ca exchange exists in vascular smooth muscles, the clear involvement of Na ions in the generation of tension can in many cases be explained by the operation of mechanisms other than Na-Ca exchange. Readers who have got this far may still be unclear whether we believe that smooth muscle cells do possess the exchange mechanism or not. In the next section we will briefly describe two more recent approaches which have convinced us that at least some smooth muscles do possess an Na-Ca exchange mechanism, one involving isolated smooth muscle membranes and the mechanisms they possess for transporting Ca, and one using ion-sensitive microelectrodes to follow in more detail changes in intracellular Na activity and membrane potential that occur when the extracellular ionic environment is manipulated.

Animals↗
PubMed1985

The effects of bicarbonate and foreign anions on chloride transport in smooth muscle of the guinea-pig vas deferens.

The selectivity of the external site of the Cl transporting mechanism in the guinea-pig vas deferens has been investigated by measurement of 36Cl uptake and efflux and by direct measurement of intracellular pH. Replacing 50% of the Cl in normal Krebs solution inhibited the 15 min uptake of 36Cl in the order NO3 greater than Br greater than SCN greater than F greater than I greater than glucuronate, both in Cl-depleted tissues and tissues pre-incubated in the 50%-Cl solutions (steady-state uptake). After 3 h incubation in these solutions, the total cellular Cl was reduced by the anions in the order Br greater than NO3 greater than I greater than SCN greater than F greater than glucuronate. Br, NO3 and I reduced the cellular Cl to less than 50% of normal, suggesting that they are actively taken up by the cells. The ability of foreign anions to inhibit a 3 min uptake of high specific activity, low concentration Cl (6.5 mM) suggests an apparent affinity series of NO3 greater than Cl = SCN = Br greater than I greater than F at the external site. Addition of NO3, Cl, Br, HCO3, F, SCN or I to a Cl-free, nominally HCO3-free bathing solution accelerated 36Cl efflux. The first four mentioned were powerful stimulants, the other three less potent. However, the exact position of HCO3 in the sequence is uncertain. The rapidity with which CO2 crosses the membrane and forms HCO3 intracellularly may allow competition between HCO3 and Cl at the internal site and so distort the result. The action of F is also questionable since this ion drastically reduces the divalent cation activity and is a metabolic inhibitor. Measurement of intracellular pH provided conclusive evidence that Cl, NO3, Br and I can exchange with HCO3 across the cell membrane. This exchange is as rapid with NO3 as with Cl but slower with Br and considerably slower with I. The results also indicate that SCN ions cross the cell membrane. It is concluded that Cl, HCO3, Br and NO3 are all translocated by the exchange carrier. I and perhaps SCN also interact with the transport mechanism, but the translocation rate is then greatly reduced. The precise order of the affinity of these anions remains uncertain but the following sequence: NO3 greater than Cl = SCN = Br greater than I greater than F is considered to be the most likely.

Animals↗
PubMed1984

Evidence for sodium-calcium exchange in the guinea-pig ureter.

The effects of high-K and low-Na solutions on the smooth muscle of the guinea-pig ureter have been examined in both normal tissues, and tissues in which the Na pump had been blocked by exposure to K-free solutions or ouabain (high-Na tissues). Tension recording, membrane potential measurements and ion analysis were used. High-K solutions depolarize normal tissues, leading to action potential generation and phasic contractions followed, at concentrations greater than 20-30 mM, by cessation of action potentials and the development of a biphasic contracture which declines slowly during continuous exposure. The contracture is abolished by Ca-antagonist drugs, procaine and Ca-free solutions. Short exposures of normal tissues to Na-free solutions do not result in tension development. Longer exposures may initiate tension, depending on the Na substitute used. Sucrose causes depolarization of the cells and spike development associated with phasic contractions, superimposed on a small contracture; Li depolarizes the cells but causes no tension generation; Tris hyperpolarizes the cells and a small increase in basal tone may be seen. On exposure to K-free solutions or ouabain, the tissues do not develop significant tone but their response to short application of high-K solutions grows with time. The tissues also develop the ability to contract on short applications of low-Na solutions. The low-Na contractures are resistant to concentrations of Ca antagonists that abolish the K responses of normal tissues, but are abolished in Ca-free solutions. The ability of the tissues to contract in Na-free solutions is accompanied by an increase in intracellular Na and loss of intracellular K. Even after several hours' exposure to ouabain, however, the tissues still contain significant amounts of K and the membrane potential is the same as, or more negative than that in normal tissues. Therefore it appears that another mechanism, apart from the Na pump, can regulate intracellular Na. On continuous exposure to Na-free solutions, the contracture declines rapidly. The decline is associated with a loss of intracellular Na. The Na-free contracture is larger when K rather than Tris is used as the substitute. This difference persists in the presence of a concentration of Mn that abolishes the K contracture of normal tissues but is abolished by high concentrations (10 mM) of procaine.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗
PubMed1984

The role of chloride-bicarbonate exchange in the regulation of intracellular chloride in guinea-pig vas deferens.

The Cl-depleted smooth muscle cells of guinea-pig vas deferens rapidly restore their intracellular Cl activity ( aiCl ) to a level 5 times higher than that predicted by a passive distribution when Cl- ions are readmitted to the extracellular solution. Cl reaccumulation, measured using Cl-sensitive micro-electrodes and the uptake of 36Cl, was slowed about 3-fold by the nominal absence of CO2 and HCO3 and about 10-fold by the presence of the anion exchange inhibitor DIDS (4-4'-diisothiocyanostilbene-2,2'-disulphonic acid). However, the usual level of intracellular Cl was eventually attained and neither condition reduced intracellular Cl in normal tissues. The loss of intracellular Cl that occurs when Cl- ions are removed from the extracellular solution was slowed about 3-fold by the nominal absence of CO2 and HCO3 and was accelerated by their readdition. DIDS slowed the fall in aiCl about 10-fold and reduced 36Cl efflux. Intracellular pH (pHi), measured using pH-sensitive micro-electrodes, increased by a mean of 0.73 units when Cl was removed from the superfusing solution in the presence of CO2 and HCO3, and rapidly decreased when Cl was readmitted. These changes are equivalent to an intracellular accumulation and loss of HCO3- ions respectively. The net transmembrane movement of HCO3- ions which would have caused these changes in pHi was calculated using the measured intracellular buffering power ( Aickin , 1984). 25% fewer HCO3- ions than Cl- ions were accumulated and lost and the HCO3 movement was completed 2-3 times faster than the simultaneous Cl movement under the same conditions. The changes in pHi induced by alteration of extracellular Cl were reduced by the nominal absence of CO2 and HCO3 and abolished by the presence of DIDS. The acidification recorded on readmission of Cl in the nominal absence of CO2 and HCO3 was compatible with a metabolic production of about 0.1% CO2. We conclude that Cl-HCO3 exchange plays a major role in the regulation of intracellular Cl. The exchange carrier is reversible, is completely inhibited by DIDS, and accounts for about 75% of the net Cl movement that occurs when Cl is removed from or readmitted to the extracellular solution. The mechanism which is responsible for the remaining Cl movement remains to be elucidated.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1983

The effects of papaverine on the electrical and mechanical activity of the guinea-pig ureter.

1. The effects of papaverine (10(-5)-2 X 10(-4) M) were studied on the evoked electrical and mechanical activity of the guinea-pig ureter smooth muscle. In normal conditions the action potential consists of an initial spike followed by further spikes superimposed on a plateau phase. Papaverine reversibly decreased the duration of the plateau of the action potential, blocked the associated spikes, greatly reduced the amplitude of the contraction but enhanced the initial component of the action potential. 2. Papaverine did not change the membrane potential and had little effect on the membrane resistance. 3. Tetraethylammonium (5 mM), which blocks the delayed outward K current, did not prevent the decrease in the duration of the plateau nor the decrease of the contractile response caused by papaverine. 4. In Na-free solution the duration of the action potential was decreased until only a single spike was seen, due to suppression of the plateau. An effect of papaverine could not be observed under these conditions. 5. Mn2+ ions (1 mM) completely suppressed the spike component and tension while the plateau component was substantially increased. Papaverine in the presence of Mn2+ reversibly blocked the generation of the action potential. When Mn2+ ions were added to Na-free solution the duration as well as the amplitude of the spike was increased. Again, papaverine reversibly blocked the generation of the action potential. 6. Noradrenaline (10(-4) M) and histamine (10(-5) M) in normal conditions prolonged the duration of the action potential plateau and increased both the duration and amplitude of the concentration. Papaverine again blocked the plateau and greatly reduced the contractile response. 7. Papaverine caused the relaxation of KCl-induced contractures, preferentially blocking the tonic component. 8. It is suggested that the inhibitory action of papaverine on ureter smooth muscle results from its specific blockade of the 'slow' Na/Ca channels responsible for the generation of the plateau component of the action potential.

Action Potentials↗
PubMed1983

Towards an estimate of chloride permeability in the smooth muscle of guinea-pig vas deferens.

Cl movements across the cell membranes of smooth muscle from the guinea-pig vas deferens were measured using Cl-sensitive micro-electrodes and 36Cl fluxes. The rate constants for the loss of Cl ions measured by both methods under a variety of conditions were used to calculate the apparent Cl permeability (PCl). If it is assumed that the initial rate of decline of the intracellular Cl activity (aiCl) on removal of extracellular Cl (Clo) represents net transmembrane Cl movement, the apparent PCl was 3-6 X 10(-8) cm s-1. This value is in good agreement with those calculated from the rate constant of 36Cl efflux into both normal Krebs solution (steady-state) and Cl-free solution. Such a value for PCl predicts a large depolarization on removal of Clo, but only a minimal change was recorded. It also predicts that changes in membrane potential (Em) would affect aiCl; furthermore that removal of Clo would increase membrane resistance and thus the hyperpolarization observed on reactivation of the electrogenic Na pump. Neither of these was observed. The PCl/PK ratio obtained from changes in Em on rapid changes in Clo and Ko gives a value for PCl which is an order of magnitude lower: 4 X 10(-9) cm s-1, using Casteels' (1969 b) value for PK. These observations can be reconciled by a substantial proportion of the measured Cl movements being carrier-mediated. The presence of the stilbene derivative DIDS greatly slowed both the steady-state efflux and uptake of 36Cl, as has previously been shown for the loss and reaccumulation of Cl ions on removal and replacement of Clo. PCl calculated in the presence of DIDS was about 5 X 10(-9) cm s-1. The nominal absence of CO2 and HCO3, which slows the reaccumulation and loss of Cl, had no effect on the steady-state fluxes. This indicates that the carrier operates in the self-exchange mode in the steady state.

Animals↗
PubMed1982

Measurement of intracellular chloride in guinea-pig vas deferens by ion analysis, 36chloride efflux and micro-electrodes.

1. Cl-sensitive micro-electrodes were used to measure the intracellular Cl activity (a(Cl) (i)) in smooth muscle cells of the guinea-pig vas deferens. The values obtained were compared with those of intracellular Cl (Cl(i)) found by both ion analysis and (36)Cl efflux.2. Various combinations of filling solution for recording membrane potential (E(m)), and type of micro-electrode were tested. The most successful, which allowed continuous recording of a(Cl) (i) for several hours, was a double-barrelled electrode using the reference liquid ion exchanger (RLIE; Thomas & Cohen, 1981). However, a(Cl) (i) measured both by simultaneous impalements of separate cells with Cl-sensitive and conventional micro-electrodes, and by double-barrelled micro-electrodes, was about 42 mM in normal Krebs solution. This is five times higher than the value from a passive distribution. E(Cl) was about -24 mV, more than 40 mV positive to E(m).3. On complete removal of extracellular Cl (Cl(o)), a(Cl) (i) fell to an apparent level of about 3 mM. If this represents interference from other anions, the maximum error in E(Cl) measured in normal Krebs solution is 2.5 mV. Replacement of Cl(o) caused a rapid increase in a(Cl) (i). This must be caused by an active transport of Cl(-) ions into the cell against their electrochemical gradient.4. The stabilized values of a(Cl) (i) measured at different levels of Cl(o) agree surprisingly well with a(Cl) (i) estimated from ion analysis and (36)Cl efflux, assuming that the intracellular activity coefficient was the same as measured in the normal Krebs solution. The relationship of a(Cl) (i) to Cl(o) was hyperbolic.5. It is concluded that Cl-sensitive micro-electrodes accurately measure a(Cl) (i) in smooth muscle cells. The remarkable agreement between the direct and indirect methods of measuring Cl(i) suggests that Cl(-) ions are not bound to a significant extent and that the compartment seen by the micro-electrodes is probably representative of the whole cell.

Animals↗
PubMed1980

A possible role of linked Na and Cl movement in active Cl uptake in smooth muscle.

Smooth muscles actively accumulate Cl. Abolition of the transmembrane Na gradient has been shown to lead to a passive distribution of Cl. This suggested to us that Na and Cl may move together across the smooth muscle cell membrane, Cl accumulation resulting due to [Na]i being less than [Na]o. In favour of this view are the findings that Cl uptake is reduced in Na-free medium, Na uptake is reduced in Cl-free medium, reducing [Cl]i reduces Na efflux, and increasing [Na]i increases Cl efflux.

Animals↗