bluuebunny/crossref_metadata_2025 huggingface dataset:
A patch‐clamp study of potassium channels and whole‐cell currents in acinar cells of the mouse lacrimal gland.
Authors: Findlay I
Published: December 2014

Individual acinar cells were isolated enzymatically from the mouse exorbital lacrimal gland. Their electrical characteristics were studied by the patch‐clamp methods of single‐channel and whole‐cell recording as described by Hamill, Marty, Neher, Sakmann & Sigworth (1981). Recording from cell‐attached and excised inside‐out patches of acinar membrane with quasi‐physiological ion gradients demonstrated large outward current events that correspond to single‐channel openings. The amplitude, frequency and duration of channel events increased as the membrane patch was depolarized and were reduced by hyperpolarization of the patch membrane. The reversal potential for these channel events is more negative than ‐40 mV. In excised inside‐out patches exposed to quasi‐physiological ion gradients single‐channel events were abolished when K+ was replaced by Rb+. Since there was no Cl‐ gradient the channel is clearly highly selective for K+. In excised inside‐out patches, when the free Ca2+ concentration bathing the physiological inside of the membrane was raised from less than 10(‐9) M to 10(‐8) M the frequency and duration of opening of the K+ channel was increased. The channel was almost continuously open when the membrane was exposed to 10(‐7) M‐free Ca2+. 'Whole cell' recording of lacrimal acinar cells containing 140 mM‐KCl and 1 mM‐EGTA (with no added Ca2+) provided cell resting membrane potentials of ‐30 to ‐40 mV. Depolarizing voltage jumps from the resting membrane potential evoked large outward currents. Hyperpolarizing voltage jumps only evoked small inward currents. Whole‐cell recording where RbCl replaced KCl in the pipette provided resting membrane potentials of ‐20 to ‐30 mV, reduced the amplitude of outward currents evoked by cell‐depolarizing voltage steps by 60% and slowed the time course of the currents. Isolated cells containing 140 mM‐KCl and 1 mM‐EGTA were voltage clamped at their resting membrane potentials. Acetylcholine (ACh) was applied locally and immediately evoked a strong outward current which rapidly declined to a steady‐state level. Sustained agonist responses were obtained by exposing the isolated cell to a solution containing 10(‐6) M‐ACh. In both K+‐ and Rb+‐filled cells, where the intracellular Ca2+ concentration was buffered by the inclusion of 1 mM‐EGTA, 10(‐6) M‐ACh evoked sustained outward currents that corresponded to cell hyperpolarizations of 5‐15 and 10‐20 mV, respectively. Increasing intracellular Ca2+ buffering by including 10 mM‐EGTA abolished secretagogue‐induced outward current in both K+‐ and Rb+‐filled cells. It is concluded that the lacrimal acinar cell membrane contains voltage‐ and Ca2+‐activated K+ channels.(ABSTRACT TRUNCATED AT 400 WORDS)
Aortic wall properties and baroreceptor behaviour at normal arterial pressure and in acute hypertensive resetting in dogs.
Authors: Coleridge H M , Coleridge J C , Poore E R , Roberts A M , Schultz H D
Published: December 2014

In order to throw light on the mechanism of acute hypertensive baroreceptor resetting, we examined the relationship between aortic baroreceptor firing and aortic wall properties in anaesthetized dogs as pressure was varied in a number of ways. We recorded baroreceptor impulses from the left aortic nerve, and measured aortic pressure with a catheter‐tip transducer and external aortic diameter with ultrasonic transit‐time transducers. Narrow anticlockwise hysteresis loops were evident in the pressure‐diameter relationship of the upper thoracic aorta, both during the rapid pulsatile pressure changes of the cardiac cycle and during the slow excursions of mean pressure imposed for construction of baroreceptor pressure‐‐response curves. In contrast to the 'phase‐lag' response of diameter to pressure, the baroreceptor response was 'phaselead' in character, decreasing when stress‐induced creep occurred in the aortic wall. When the mean arterial pressure set‐point was increased from 100 to 125 mmHg for 20 min, the hysteresis loops relating mean diameter to mean pressure in the range 60‐200 mmHg were displaced along the diameter axis in the direction of wall creep. A reduction in the baroreceptor response to pressure (i.e. resetting) always accompanied this displacement. Administration of ouabain (25‐35 micrograms/kg) had no consistent effect on baroreceptor resetting. It has been suggested that acute baroreceptor resetting is akin to adaptation. To investigate the possibility that the two processes are accompanied by similar changes in aortic wall properties, we converted the aorta into a closed sac and distended it with a square wave of pressure. Like resetting, adaptation of the baroreceptor response to maintained pressure was associated with a small degree of creep of the aortic wall. Our results are compatible with the hypothesis that acute hypertensive resetting of aortic baroreceptors is similar to adaptation, both phenomena being attributable to relaxation of viscoelastic coupling elements, leading to a reduction of strain at the receptor membrane. Whether viscoelastic processes alone can account for acute resetting, or whether changes in ionic balance are involved also, baroreceptor responsiveness is a function of the stress history of the wall, the pressure‐response curve moving along the pressure axis in the direction of the prevailing set‐point. Hence, in early hypertension physiological resetting of baroreceptors will precede pathological resetting, and may even promote an upward movement of set‐point.
The ventilatory response of the human respiratory system to sine waves of alveolar carbon dioxide and hypoxia.
Authors: Robbins P A
Published: December 2014

Sine waves of alveolar CO2 at constant high alveolar O2, and sine waves of alveolar hypoxia (1/(PA, O2 ‐C), C congruent to 32 torr) at constant alveolar CO2 have been administered to three subjects in each case. Sine waves of six different periods were studied, ranging between 900 and 30 s for the CO2 sine waves and 300 and 20 s for the hypoxic sine waves. The sinusoidal variations in ventilation produced by these manoeuvres, expressed as amplitudes and phase shifts, were compared with values predicted from the dynamic responses to alveolar steps of gas tension already to be found in the literature. For the CO2 sine waves, the amplitudes of response agreed well with those predicted at the higher frequencies, but were less than predicted at the lower frequencies. For the hypoxic sine waves, the amplitude of response varied less with frequency than was predicted. For both the CO2 and the hypoxic sine waves, the phase shift of response was less than expected at the higher frequencies. An attempt was made to fit parameters to a simple model, based on the wash‐in and wash‐out of respiratory gases into and out of a tissue compartment, and used in the literature for describing the responses to step changes. No satisfactory fit was found. It is concluded that the simple model is unsatisfactory by itself for describing the responses to sinusoidal chemical stimulation; features additional to those included in the model are required to explain fully the responses seen. The possibilities for chemoreception at the higher frequencies are discussed in the light of the low phase shifts.
Factors influencing free intracellular calcium concentration in quiescent ferret ventricular muscle.
Authors: Allen D G , Eisner D A , Orchard C H
Published: December 2014

The photoprotein aequorin was injected into cells of ferret papillary muscles to monitor the resting intracellular free Ca concentration [( Ca2+]i). Increasing the external Ca concentration [( Ca2+]o) increased both resting [Ca2+]i and resting tension. The tension and [Ca2+]i both rose to a peak and then declined to a steady‐state level which was higher than the control. Qualitatively similar, but larger, effects were observed if [Ca2+]i was first elevated with strophanthidin. The increase of [Ca2+]i was accompanied by the development of spontaneous oscillations of [Ca2+]i. When a steady level of [Ca2+]i had been reached in high [Ca2+]o, [Ca2+]o was reduced back to the control level for a brief period. A subsequent increase of [Ca2+]o produced a rise of [Ca2+]i to the same steady level as that previously found in the high [Ca2+]o but the initial peak and subsequent decline were absent. It is suggested that the decline of [Ca2+]i from the initial peak is mediated by a fall of intracellular Na concentration [( Na+]i) limiting Ca entry on a Na‐Ca exchange. Increasing external K concentration [( K+]o) from 5 to 30 mmol/l had no detectable effect on [Ca2+]i under control conditions. However, if [Ca2+]i was first increased either by applying strophanthidin or by increasing [Ca2+]o, increasing [K+]o produced a transient rise of [Ca2+]i and tension. This rise was unaffected by D600. It is suggested that the secondary decline of [Ca2+]i after the initial rise may, again, be produced by a fall of [Na+]i acting on an Na‐Ca exchange. Acidification produced by increasing [CO2] had no detectable effect on [Ca2+]i under control conditions. However, if [Ca2+]i was increased by strophanthidin, acidification produced a rise of [Ca2+]i. This rise of [Ca2+]i was partly transient even when the intracellular acidification was presumably maintained (raising CO2 at constant [HCO3‐]). Acidification in Na‐free solutions had qualitatively similar effects to those in Na‐containing solutions. In Na‐free solutions (Na replaced by K) the [Ca2+]i could be maintained at a low level for at least several hours. Increases of [Ca2+]o in Na‐free solutions led to a decrease of [Ca2+]i, and similarly decreasing [Ca2+]o led to an increase in [Ca2+]i. These anomalous effects of [Ca2+]o on [Ca2+]i could be abolished by Mn ions or D600. It is suggested that changes in [Ca2+]o may have reciprocal effects on Ca permeability and hence on [Ca2+]i. The application of the mitochondrial uncoupler FCCP in Na‐free solutions led to an increase of resting tension followed, after a substantial delay, by an increase of [Ca2+]i.(ABSTRACT TRUNCATED AT 400 WORDS)
Effect of sialoadenectomy and salivary gland extracts on gastrointestinal mucosal growth and gastrin levels in the rat.
Authors: Skinner K A , Soper B D , Tepperman B L
Published: December 2014

We have examined gastrointestinal mucosal growth 30 days after surgical removal of the submandibular‐sublingual salivary glands and ligation of the parotid ducts of rats. The rate of [3H]thymidine uptake in vitro as an estimation of DNA synthesis and the content of DNA and RNA were examined in the oxyntic, duodenal and proximal colonic mucosa. DNA synthesis, DNA and RNA content of oxyntic mucosa were reduced in sialoadenectomized rats when compared to sham‐sialoadenectomized control animals. There was no change in the degree of [3H]thymidine incorporation or DNA content of the duodenal or colonic mucosa. Intraperitoneal injection of an aqueous extract of the submandibular‐sublingual salivary glands of rats (4.0 mg tissue protein in 0.1 M‐sodium phosphate buffer administered twice a day for 15 days) increased the rate of DNA synthesis and the total mucosal DNA and RNA content in the oxyntic mucosa. Injections of extracts of spleen or muscle did not produce consistent results. Administration of epidermal growth factor (10 micrograms/kg) or pentagastrin (250 micrograms/kg) resulted in an increase of the level of DNA synthesis observed in the oxyntic mucosa of sialoadenectomized rats. Plasma and antral tissue levels of the trophic hormone, gastrin, were not significantly decreased in sialoadenectomized rats treated with 0.1 M‐sodium phosphate‐buffered saline. However, treatment with the salivary tissue extract did result in significant reductions in both plasma and tissue levels of gastrin. We conclude that elimination of the major salivary glands in the rat results in a decrease in [3H]thymidine uptake and DNA content of the gastric oxyntic mucosa. These effects are not mediated via a reduction in endogenous levels of the trophic hormone, gastrin. Administration of an aqueous extract of salivary tissue exerted a small but significant trophic influence on the oxyntic mucosa of the rat.
Pre‐and post‐junctional effects of tubocurarine and other nicotinic antagonists during repetitive stimulation in the rat.
Authors: Gibb A J , Marshall I G
Published: December 2014

The effects of tubocurarine and trimetaphan have been examined at voltage‐clamped rat diaphragm neuromuscular junctions during (a) single and repetitive stimulation of the phrenic nerve in cut muscles and (b) repetitive ionophoretic application of acetylcholine (ACh). Tubocurarine (2.5 X 10(‐7)‐10(‐6)M) produced a concentration‐dependent reduction in the amplitude of neurally evoked end‐plate currents (e.p.c.s). It also reduced their time constant of decay (tau e.p.c.) in a manner that was independent of membrane potential, and not markedly dependent on the tubocurarine concentration. Likewise the snake alpha‐neurotoxin, erabutoxin b, reduced the e.p.c. amplitude and produced a voltage‐independent shortening of tau e.p.c. Estimates of mean channel lifetime (tau noise) from ACh‐induced e.p.c. fluctuations revealed that (a) tau noise was 46.4 +/‐ 3.7% shorter than tau e.p.c. measured at the same end‐plate. At these same end‐plates in the presence of tubocurarine (2.5 X 10(‐7)M) tau e.p.c. was 32.6 +/‐ 1.0% shorter than the control tau e.p.c. but tubocurarine did not change tau noise, (b) trimetaphan (2.5 X 10(‐5)‐2 X 10(‐4)M) produced a concentration‐dependent and voltage‐dependent reduction of tau e.p.c., and a concentration‐dependent reduction of peak e.p.c. amplitude. Trimetaphan (2.5 X 10(‐5)M) produced a 50% reduction of tau noise. (a) Both tubocurarine and trimetaphan produced concentration‐dependent increases in the run‐down of trains of neurally evoked e.p.c.s (50 Hz, 0.4 s). This effect did not vary with membrane potential in tubocurarine, but was voltage dependent when induced by trimetaphan. (b) Erabutoxin b reduced the e.p.c. amplitude but did not produce any increase in the run‐down of trains of neurally evoked e.p.c.s. During 50 Hz repetitive ionophoretic application of ACh, tubocurarine (2.5 X 10(‐7)M) reduced the amplitude of each current in the train without inducing any run‐down of the current amplitudes. This effect was not dependent on the membrane potential. In contrast trimetaphan (2.5 X 10(‐5)M) induced a voltage‐dependent run‐down of trains of ionophoretically evoked e.p.c.s. We conclude that tubocurarine and erabutoxin b reduce the e.p.c. amplitude by blocking the post‐junctional ACh receptor. Tubocurarine produces tetanic rundown of e.p.c.s. by a prejunctional mechanism, whereas the effects of trimetaphan during single and repetitive stimulation are at least partly due to block of the open ion channel associated with the ACh receptor.
The reversal potential of excitatory amino acid action on granule cells of the rat dentate gyrus.
Authors: Crunelli V , Forda S , Kelly J S
Published: December 2014

The responses of granule cells to glutamate, aspartate, N‐methyl‐D‐aspartate (NMDA), quisqualate and kainate applied by ionophoresis on to their dendrites in the middle molecular layer of the dentate gyrus were studied with intracellular electrodes using an in vitro hippocampal slice preparation. On passive depolarization 75% of the granule cells displayed anomalous rectification, which persisted in the presence of TTX and TEA but was eliminated by Co2+ or the intracellular injection of Cs+. Short ionophoretic applications of all the excitatory amino acids evoked dose‐dependent depolarizations that were highly localized: movement of the ionophoretic electrode by as little as 10 microns could substantially change the size of the response. The depolarizations evoked by glutamate, asparatate, quisqualate and kainate were unaffected by TTX and Co2+. The depolarization evoked by NMDA was unaffected by TTX but markedly reduced by Co2+. Following intracellular injection of Cs+, neurones could be depolarized to +30 mV and the depolarizations produced by glutamate, quisqualate, NMDA and kainate reversed. The reversal potentials (E) were Eglutamate: ‐5.6 +/‐ 0.4 mV; ENMDA: 1.8 +/‐ 1.9 mV; Equisqualate: ‐3.9 +/‐ 1.9 mV; Ekainate: ‐4.6 +/‐ 2.0 mV. The excitatory post‐synaptic potential (e.p.s.p.) evoked by stimulation of the medial perforant path could also be reversed and Ee.p.s.p. was ‐5.5 +/‐ 1.1 mV. The 6 mV difference between ENMDA and the equilibrium potential for the other exogenously applied excitatory amino acids and the statistically significant difference between ENMDA and Ee.p.s.p. (P less than 0.005; d.f.: 7) is consistent with our earlier hypothesis that both the transmitter released by the medial perforant path and exogenously applied glutamate are unlikely to interact with NMDA receptors.
Slow excitatory synaptic potentials recorded from neurones of guinea‐pig submucous plexus.
Authors: Surprenant A
Published: December 2014

Intracellular recordings made from neurones of guinea‐pig submucous plexus revealed three types of synaptic input: cholinergic excitatory synaptic potentials (fast e.p.s.p.s) of 50‐80 ms duration, inhibitory synaptic potentials (i.p.s.p.s) of 1 s duration, and non‐cholinergic, non‐adrenergic slow e.p.s.p.s which lasted for 15‐20 s. A single stimulus was sufficient to elicit the slow e.p.s.p. in all neurones in which this synaptic input was present. Slow e.p.s.p.s were recorded in those neurones which also displayed i.p.s.p.s. Both the i.p.s.p. and the slow e.p.s.p. appeared in an all‐or‐none fashion and were not affected by alterations in the stimulus strength. The inhibitory as well as the slow excitatory synaptic potentials reversed close to the K+ equilibrium potential, indicating that the i.p.s.p. is due to an activation of K+ conductance while the slow e.p.s.p. is due to its inactivation. Evidence is presented which suggests the slow e.p.s.p. may be generated at a synapse located some distance from the soma, presumably at a dendritic location. Only those cells which showed slow e.p.s.p.s responded to substance P with a depolarization which mimicked the slow e.p.s.p.
Two types of neurones lacking synaptic input in the submucous plexus of guinea‐pig small intestine.
Authors: Surprenant A
Published: December 2014

Intracellular recordings were made from neurones in the submucous plexus of guinea‐pig small intestine. Approximately 20% of all neurones studied appeared to lack synaptic input; two types of neurones without synaptic input were identified. The action potential recorded from one of these types of ganglion cells was followed by a prolonged after‐hyperpolarization. The after‐hyperpolarization was abolished by Co2+ or Mn2+. A Ca2+ component of the evoked action potential was demonstrated after the Na+ component had been abolished by tetrodotoxin (TTX). The other type of neurone was characterized by a high input resistance (220‐320 M omega) and spontaneous action potentials which were unaffected by hexamethonium, curare, atropine or TTX. The ionic basis of the action potentials evoked from spontaneously active cells differed from other enteric neurones. These action potentials exhibited a TTX‐insensitive Na+ component as well as a Ca2+ component. It is suggested the spontaneously active neurones may be responsible for the ongoing synaptic activity recorded from many submucous plexus neurones in the absence of stimulation.
Comparison of cholinergic activation and desensitization at snake twitch and slow muscle fibre end‐plates.
Authors: Connor E A , Fiekers J F , Neel D S , Parsons R L , Schnitzler R M
Published: December 2014

Characteristics of receptor‐channel activation and desensitization have been compared at voltage‐clamped snake slow and twitch fibre end‐plates maintained in an isotonic potassium propionate solution. Miniature end‐plate current (m.e.p.c.) decay was slower and less voltage dependent at slow fibre end‐plates than at twitch fibre end‐plates. The peak m.e.p.c. amplitude versus voltage relationship and reversal potential were similar at the two end‐plate types. Acetylcholine‐induced noise and m.e.p.c.s were recorded at slow fibre end‐plates. At most slow fibres the spectral density was not adequately fitted by a single Lorentzian function. Rather, the observed spectral density was greater at high frequencies than the values predicted using the m.e.p.c. decay rate. The noise could be well described by the sum of two Lorentzian functions, one of which corresponded to a single Lorentzian function with the corner frequency determined by the m.e.p.c. decay rate. The shape of the carbachol concentration‐peak end‐plate current relationship was similar at both slow and twitch fibre end‐plates. However, for all concentrations tested, the peak carbachol‐induced end‐plate current (e.p.c.carb.) value was markedly less at slow fibre end‐plates than at twitch fibre end‐plates. The onset of desensitization was determined using two methods. The first concerned analysis of the time course of decay of the e.p.c.carb. from a peak value during the sustained application of agonist. The second involved a double‐perfusion technique in which a 'desensitizing' dose was applied for varying intervals before the application of a second 'test' dose of carbachol. With both methods the development of desensitization at both end‐plate types was dependent on carbachol concentration and duration of exposure. At each end‐plate type the time course of desensitization onset often exhibited two components; one with a time constant of seconds and a slower component having time constants in the range of tens to hundreds of seconds. The slope of the relationship between carbachol concentration and equilibrium desensitization at slow and twitch fibre end‐plates was close to two, suggesting that two molecules of agonist are probably bound during the development of desensitization. However, for all concentrations tested, desensitization developed more rapidly and to a greater extent at twitch fibre end‐plates than at slow fibre end‐plates. The voltage dependence of the 3 min steady‐state desensitization produced by 108 microM‐carbachol was very similar (approximately ‐0.0250 mV‐1) at both fibre types.(ABSTRACT TRUNCATED AT 400 WORDS)
Generation of an unusual depolarizing response in rabbit primary afferent neurones in the absence of divalent cations.
Authors: Stansfeld C E , Wallis D I
Published: December 2014

The effects of divalent cations on responses to 5‐hydroxytryptamine (5‐HT), gamma‐aminobutyric acid (GABA) and 1,1‐dimethyl‐4‐phenyl piperazinium (DMPP) were investigated using a sucrose‐gap method to record population responses. In Ca‐free medium responses to 5‐HT were enhanced, those to DMPP depressed and those to GABA unchanged. In Mg‐free medium responses to 5‐HT were unchanged, while those to DMPP and GABA were depressed. Removal of both Ca and Mg from the superfusion medium caused a small reduction of GABA responses and a large reduction of DMPP responses. Responses to 5‐HT were not only greatly potentiated but were changed in character; the depolarizing phase became sigmoid and the dose dependence between quantity of 5‐HT and response magnitude was lost as if 5‐HT were triggering an all‐or‐nothing phenomenon. Dose‐‐response relationships for GABA were normal in the large majority of preparations. In about 10% of preparations, supramaximal amounts of GABA or DMPP evoked large responses of a similar character to those evoked by 5‐HT. The large responses, generated by an unknown mechanism, were termed X responses. Further reduction in tissue divalent cations by EGTA (1 mM) caused X responses to be generated spontaneously. Ca, Mg, Mn or Co (1 mM) could suppress X responses. DMPP responses, reduced in Ca/Mg‐free medium, were largely restored by 1 mM‐Ca. Depression of GABA responses in Ca/Mg‐free medium could be entirely attributed to the absence of Mg, Mn being able to substitute for Mg. X responses were generated only after equilibration for 1 h with Ca/Mg‐free medium. Attempts to manipulate [Ca]i with dinitrophenol or caffeine did not produce the conditions under which X responses were generated. Intracellular records of responses to 5‐HT, GABA or DMPP showed that cells with A fibres responded to GABA but not to 5‐HT or DMPP. Fifty‐four out of sixty‐seven cells with C fibre axons (80%) were depolarized by 5‐HT, thirty‐seven out of forty‐nine (76%) by DMPP and forty out of fifty‐seven (70%) by GABA. Eighteen out of thirty‐eight (47%) C cells were depolarized by all three agents. Some C cells were very sensitive to 5‐HT, 10(‐6) M evoking a substantial response. In most, responses to 10(‐5) M‐5‐HT had a slower rate of rise than responses to 10(‐4) or 10(‐3) M‐GABA or DMPP, yet lower 5‐HT concentrations normally elicited X responses in sucrose‐gap experiments whereas GABA or DMPP normally did not.(ABSTRACT TRUNCATED AT 400 WORDS)
Xenopus oocyte resting potential, muscarinic responses and the role of calcium and guanosine 3',5'‐cyclic monophosphate.
Authors: Dascal N , Landau E M , Lass Y
Published: December 2014

Resting potential (r.p.) and muscarinic response mechanisms were studied in Xenopus laevis oocytes using the voltage‐clamp technique. Insertion of micro‐electrodes into the oocyte produced a 'shunt' membrane conductance which partially sealed after a few minutes. The oocyte resting potential (measured with a single intracellular electrode) ranged from ‐40 to ‐60 mV. Ouabain and low K+ solution depolarized both follicles and denuded oocytes. The electrogenic Na+‐K+ pump was more active in the latter. In the presence of ouabain, the r.p. agreed with the constant field theory. alpha (PNa+/PK+) was 0.12 in follicles and 0.24 in denuded oocytes. beta (PCl‐/PK+) was 0.4 in both. At [Na+]o lower than 70 mM, the r.p. deviated considerably from the constant field predictions. The relatively large value of alpha indicated the major role of Na+ in oocyte r.p. determination. The oocyte muscarinic response was separated into four distinct components: the fast depolarizing Cl‐ current, 'D1'; the slow depolarizing Cl‐ current, 'D2'; the slow hyperpolarizing K+ current, 'H'; and the large membrane Cl‐ current fluctuation, 'F'. The H response reversal potential showed a Nernst relationship to [K+] and was selectively blocked by intracellular injection of tetraethylammonium (TEA). The D1 and D2 reversal potential showed a Nernst relationship to [Cl‐]. In Ca2+‐deficient, EGTA‐containing medium, D2 and F were abolished and D1 and H were reduced. Verapamil inhibited all responses. Increasing [Ca2+]o caused a significant increase in D1, D2 and F response amplitudes. Intracellular injection of 0.6‐10 pmol guanosine 3',5'‐cyclic monophosphate, induced a large outward K+ current, similar to the muscarinic H response.
Characteristics of ionic binding by rat renal tissue in vitro.
Authors: Law R O
Published: December 2014

A study has been made of Na+ and Cl‐ binding in metabolically inhibited slices of rat renal cortex and outer medulla incubated in modified Krebs phosphate‐bicarbonate Ringer solution. At pH 7.35 in control media (cortex, 147 mmol Na+/l, 105 mmol Cl‐/l; outer medulla, 187 mmol Na+/l, 145 mmol Cl‐/l) cortical slices bound (mean) 171 nmol Na+ and 56.7 nmol Cl‐/mg solute‐free dry weight; outer medullary slices bound 188 nmol Na+/mg and negligible amounts of Cl‐. In both regions, Na+ was exchangeable on a 1:1 basis for K+ or Li+ in media containing equal concentrations of each cation: Na+ was completely displaced by La3+. In cortical slices in media containing equimolar Cl‐ and other monovalent anions, binding occurred according to the sequence acetate less than or equal to salicylate less than or equal to Cl‐ less than SCN‐; Cl‐ was completely displaced by PO4(3‐). When medium pH was lowered, Na+ binding was markedly reduced in both regions, whereas Cl‐ binding increased (and became significant in outer medulla). In NaCl solutions, Na+ binding capacity was saturated at control Na+ concentrations. When [Na+] was progressively reduced (iso‐osmolality being maintained by addition of urea), bound Na+ in both regions was nearly linearly related to log medium [Na+]. Raising medium osmolality with urea caused decreased Na+ binding and increased Cl‐ binding in both regions. Na+ binding in both regions was significantly reduced by pre‐treatment with chondroitinase ABC. Binding of both ions was independent of temperature within the range 2‐37 degrees C. The possibility is raised that renal ionic binding might influence vectorial ion transport by affecting free ion activity in the region of the transporting cells.
Intracellular calcium measured with calcium‐sensitive micro‐electrodes and Arsenazo III in voltage‐clamped Aplysia neurones.
Authors: Gorman A L , Levy S , Nasi E , Tillotson D
Published: December 2014

Selected neurones of the abdominal ganglion of Aplysia californica were voltage clamped, injected with the Ca2+‐indicator dye Arsenazo III, and impaled with Ca2+‐selective micro‐electrodes. Measurements of the absorbance signal (Arsenazo III) and Ca2+ micro‐electrode potential during and following voltage‐dependent Ca2+ influx (induced by voltage‐clamp pulses) were simultaneously recorded. In neurones held at ‐50 mV, the mean intracellular free Ca2+ concentration [( Ca]i) measured by the Ca2+ micro‐electrode was 0.18 microM, S.D. = 0.22 microM, n = 13. Bathing the cell in 0 Ca2+ artificial sea water (ASW) or intracellularly injecting EGTA decreased the resting [Ca]i. Voltage‐clamp pulses, which maximally activated Ca2+ channels (from ‐50 to +30 mV), transiently increased both the Arsenazo III absorbance and the Ca2+ micro‐electrode signals, indicating a rise in [Ca]i. Given the Ca2+ micro‐electrode's limited band width, the peak of the Ca2+ signal during the pulse train could not be resolved; however, there was a net deflexion of this signal following the last pulse which slowly decayed to base line. Bathing the cells in 0 Ca2+ ASW, or reducing the driving force for Ca2+ entry (by stepping the voltage‐clamp pulses to much higher membrane potentials) dramatically reduced both the absorbance and the Ca2+ micro‐electrode signal increases. On the other hand, bathing the cells in 100 mM‐Ca2+ ASW increased both signals. The intracellular Ca2+ gradient within the cytoplasm following voltage‐clamp pulses was investigated by moving the Ca2+‐selective micro‐electrode tip in a step‐wise manner relative to the membrane surface. The measured rise in [Ca]i was greatest near the membrane and not measurable within 40‐50 microns of the membrane surface. The amplitude of the [Ca]i rise at different distances from the membrane could be fitted by a model based on a simple diffusion of Ca2+ from a plane source.
Experimental analysis of the relationship between charge movement components in skeletal muscle of Rana temporaria.
Authors: Adrian R H , Huang C L
Published: December 2014

Experiments were performed to ascertain whether the monotonic (q beta) and delayed (q gamma) components of non‐linear charge in skeletal muscle membranes form a sequential system, or are the result of separate, independent processes. The non‐linear capacitance studied in a large number of fibres increased with fibre diameter. This dependence was attributable to tetracaine‐sensitive (q gamma) but not to tetracaine‐resistant (q beta and q alpha) charge. The kinetics and total quantity of q gamma charge moving in response to voltage steps from varying pre‐pulse potentials to a fixed probe potential remained constant despite variations in the size of the early q beta decay. The kinetics of the delayed (q gamma) charging current obtained from a single 20 mV depolarizing step were compared with the sum of the responses to two 10 mV steps adding to the same voltage excursion. The respective transients superimposed only if one of the 10 mV steps did not reach the voltage at which q gamma first appears. In the two preceding experiments, total charge was conserved. These results are consistent with separate and functionally independent q beta and q gamma systems of potential‐dependent charge, with q gamma residing in the transverse tubules and q beta on surface membrane. The findings can be discussed in terms of a contractile 'activator' with a steep sensitivity to voltage that begins only with depolarization beyond a level close to the actual mechanical threshold.
Developmental aspects of steroid‐induced ammonia release from isolated sections of rat intestine.
Authors: Coates D R , Snart R S
Published: December 2014

Release of ammonia from isolated intestinal sections of adult male rats is higher than that measured using immature animals. The increase appears to be Na+ dependent and develops during the spurt of growth at puberty. Developmental changes in Na+‐dependent ammonia release from isolated sections of the intestine and growth of the small intestine in male and female rats have been compared. Intestinal growth increases far more rapidly than body weight and in the males critical developmental changes occur early during weaning and during puberty. In females the major change is at weaning and little further change occurs during puberty. Treatment of young animals with aldosterone or testosterone increases the Na+‐dependent ammonia release precociously. Dose‐response effects of testosterone and aldosterone in distal sections of the small intestine have been compared.
The intramural pelvic nerves in the colon of dogs.
Authors: Fukai K , Fukuda H
Published: December 2014

The intramural pelvic nerves of the dog colon were studied morphologically and electrophysiologically. These nerves emanate from the pelvic plexus, and ascend between the muscle layers along 57.5% of the colon's length. Many thin branches connect with Auerbach's plexus. Most of the nerves responded bilaterally with compound action potentials of A delta and C fibres to stimulation of the sacral dorsal roots, and with those of C fibres to stimulation of the ventral roots. Mean conduction velocities of the A delta and C fibres were 9.0 and 0.9 m/s, respectively. Distension of a section of colon at up to 85.0% of the length of the colon from the anus activated afferent fibres, and electrical stimulation of points at up to 102% of this length elicited responses in efferent fibres. In dogs in which the continuity of the colonic wall was interrupted by a ligature but the nerves were left intact, the intramural pelvic nerves were found to convey centrifugal activities of the recto‐ and ano‐colonic reflexes to the proximal colon, and centripetal and centrifugal activities of the colo‐colonic reflex.
Location and function of voltage‐sensitive conductances in retinal rods of the salamander, Ambystoma tigrinum.
Authors: Baylor D A , Matthews G , Nunn B J
Published: December 2014

The functional role and spatial location of voltage‐sensitive conductances that modify the light‐evoked electrical signals were studied in retinal rods of the tiger salamander. An isolated rod was drawn into a suction electrode for recording membrane current and impaled with an intracellular electrode for recording membrane potential and passing current. A bright flash gave a rapid initial hyperpolarization that relaxed to a smaller plateau. Simultaneously the dark current of the outer segment was shut off with the time course of a rounded step function. This characteristic difference between the wave forms demonstrates that the voltage relaxation does not result from reopening of light‐sensitive channels. The voltage relaxation in (2) did not require light or interruption of the dark current, as the wave form was duplicated by suddenly switching off a depolarizing current injected during steady saturating light. This is explained if the relaxation depends purely on voltage‐sensitive conductances. The voltage response to a dim flash reached its peak value before the current response. The voltage wave form was predicted assuming that the recorded photo‐current drove a linear high‐pass filter with parameters derived from analysis of the voltage response to injection of a current step. When the intracellular voltage was changed by current injection the slope resistance of the outer segment slowly declined to a lower level, indicating that the outer segment contains a voltage‐sensitive conductance. When a current step was injected in bright steady light, the current recorded from the outer segment consisted of a capacity component proportional to dV/dt and a small extracellular leakage current but no detectable ionic current. This supports other evidence indicating that light‐sensitive channels comprise the main or exclusive ionic conductance of the outer segment. The behaviour in (5) is explained if the light‐sensitive channels themselves are slowly opened by hyperpolarization and closed by depolarization. Analysis of the current‐injection experiments suggests that most of the high‐pass filtering in a rod results from the action of voltage‐sensitive conductances located in the inner segment. Addition of 10 mM‐CsCl to the Ringer solution abolished the relaxation in the voltage response to a bright flash but left intact the high‐pass filtering of small signals. This would be explained by a selective block of one of two sets of voltage‐sensitive channels in the inner segment or by a voltage‐sensitive block of one kind of channel.
Developmental changes in intra‐enterocyte cation activities in hamster terminal ileum.
Authors: Cremaschi D , James P S , Meyer G , Rossetti C , Smith M W
Published: December 2014

Intracellular Na+ and K+ activities have been determined under visual control in enterocytes located at known positions along hamster ileal villi in vitro. The intracellular K+ activity found in enterocytes migrating over the basal third of villi was significantly less than that found in older enterocytes (83 and 113 mM respectively). The intracellular Na+ activity remained unaffected by the state of enterocyte development (32 mM throughout). Extracellular K+ and Na+ activities measured close to the luminal surface of the intestine were always similar to those measured in the bulk of the bathing medium. There was, however, a small difference in K+ activity detected between the bulk solution and solution near the luminal surface of enterocytes (3 . 4 and 3 . 6‐3 . 7 mM respectively). Na+ activity in the villus core was also similar to that of the bathing medium. The brush border membrane content of sucrase‐isomaltase, determined cytochemically, was found to increase rapidly as enterocytes migrated over the basal third of the villus and to plateau at a point roughly equivalent to that where intracellular K+ activities became maximal. The numerous ways by which enterocytes might control their intracellular K+ concentrations during development are emphasized. Attention is nevertheless drawn to the close correspondence seen between different aspects of brush border membrane development and the ability of enterocytes to maintain high internal concentrations of K+.
The diverse effects of noradrenaline and other stimulants on 86Rb and 42K efflux in rabbit and guinea‐pig arterial muscle.
Authors: Bolton T B , Clapp L H
Published: December 2014

The effects of noradrenaline and of raised external potassium ([K+]o) on the efflux of 86Rb or 42K and on tension were studied in preparations taken from eight different arteries under various conditions. There was a 10‐fold variation in the maximum 86Rb efflux evoked by noradrenaline (10(‐5)‐10(‐4) M) in the arteries studied, even though tension generated was comparable. Arterial contractions were either accompanied by large increases in 86Rb efflux, e.g. rabbit ear artery and aorta, guinea‐pig and rabbit pulmonary artery, or by small increases, e.g. rabbit and guinea‐pig mesenteric artery, rabbit brachial artery and guinea‐pig abdominal aorta. Raising [K+]o also had a diverse effect on 86Rb and 42K efflux: arteries giving small increases in efflux to noradrenaline also gave small increases in efflux to raised [K+]o. The maximum efflux evoked by raised [K+]o was on average three times greater than the maximum efflux evoked by noradrenaline in the arteries studied. The heterogeneity of the efflux response could not be explained by the quantitative heterogeneity of the efflux response could not be explained by the quantitative differences in the effects of noradrenaline or of raised [K+]o on membrane potential or, in the case of noradrenaline, by differences in the alpha‐receptors. In arteries in which the noradrenaline‐evoked 86Rb efflux was small, histamine, 5‐hydroxytryptamine, vasopressin and angiotensin also had little effect. Conversely, where noradrenaline produced a large increase in 86Rb efflux those other stimulants had comparable effects. Removal of extracellular calcium only slightly reduced the increment in 86Rb efflux evoked by 66 mM‐external K+ in the rabbit aorta even though contractions were virtually abolished under these conditions. In the case of 10(‐5) M‐noradrenaline, 40% of the contraction remained and its effect on efflux was significantly increased (P less than 0.05) in calcium‐free conditions. Essentially similar results were obtained using 42K. Tetraethylammonium (10‐20 mM) produced a significant and substantial reduction (P less than 0.001) in the 86Rb efflux evoked by raised [K+]o while only slightly affecting the noradrenaline‐evoked efflux in the rabbit aorta. It was concluded from these efflux experiments on vascular muscle that the channels through which potassium can escape, opened by depolarization and by activation of alpha‐receptors with noradrenaline, are from different populations, and that their properties vary from one artery to another. We have been unable to detect any substantial calcium‐activated component in 42K or 86Rb efflux responses to raised [K+]o or to noradrenaline.
The effect of calcium ions on the glutamate response and its desensitization in crayfish muscle fibres.
Authors: Thieffry M
Published: December 2014

The responses of crayfish muscle fibres to bath application or long ionophoresis of L‐glutamate were studied in normal and low Ca2+ solutions. The smaller responses recorded in low Ca2+ solutions have characteristics suggesting a faster desensitization. Desensitization and recovery have complex kinetics. Desensitization is faster and recovery slower when external Ca2+ concentration is reduced. Both components of the recovery phase, which can be fitted by the sum of two exponentials, are affected by the external Ca2+ concentration. Recovery can be accelerated by external Ca2+ ionophoresis onto desensitized glutamate receptors. Responses to brief glutamate pulses of low intensity are not affected by Ca2+ reduction. For higher intensities, signs of desensitization are detectable early in the rising phase of the response. Concanavalin A (Con A) blocks both desensitization and Ca2+ dependence with similar time courses. Whether or not the preparation has been treated with Con A, the slowly rising responses recorded in isotonic Ca2+ do not show signs of desensitization. Con A causes a partial blockade of the glutamate response. The Ca2+ dependence of the glutamate response can be explained by the Ca2+ dependence of the desensitization process, the cation acting at ectocellular sites of the muscle membrane.
The release of gamma‐aminobutyric acid from horizontal cells of the goldfish (Carassius auratus) retina.
Authors: Ayoub G S , Lam D M
Published: December 2014

Isolated horizontal cells from goldfish retinas were prepared by enzymatic dissociation using papain and separated from other cells by velocity sedimentation. In the intact retina, H1 horizontal cells possess a high‐affinity mechanism for accumulating gamma‐aminobutyric acid (GABA). This property is retained in isolated cells, which also release the accumulated GABA in response to depolarization by elevated external K+. L‐Glutamic acid and its analogues are highly effective at micromolar concentrations in eliciting the release of preloaded GABA from isolated cells. At saturating concentrations, L‐aspartic acid stimulates about one‐third as much release as L‐glutamic acid. In contrast, the D‐isomers of glutamate and aspartate are ineffective. In the intact retina, micromolar concentrations of L‐glutamic acid analogues are also capable of eliciting GABA release from H1 horizontal cells. Release of the accumulated GABA from isolated H1 cells is largely independent of external Ca2+ concentrations. In the intact retina, H1 horizontal cells also possess a K+‐stimulated GABA release mechanism that is independent of the Ca2+ concentrations in the medium. In addition, there appears to be a small but significant amount of [3H]GABA release that may be Ca2+ dependent. Under our conditions, [3H]GABA release from isolated cells is unaffected by external Na+ concentrations between 20 and 120 mM. However, concentrations of 10 mM or less significantly diminishes this release, with 70% curtailed in Na+‐free solutions. Our results, together with morphological observations by a number of other investigators, suggest that there may be two distinct mechanisms for GABA release from goldfish H1 horizontal cells: one being a conventional vesicular mechanism which is Ca2+ dependent, while the other is Na+ driven and Ca2+ independent. H1 horizontal cells in the intact goldfish retina release the accumulated GABA in response to brief incubations in darkness, which is known to be the natural stimulus that depolarizes these neurones.
The quantitative relationship between twitch tension and intracellular sodium activity in sheep cardiac Purkinje fibres.
Authors: Eisner D A , Lederer W J , Vaughan-Jones R D
Published: December 2014

Tension was measured in voltage clamped sheep cardiac Purkinje fibres while simultaneously measuring the intracellular Na activity (aiNa) with a recessed‐tip, Na‐selective micro‐electrode. Inhibiting the Na‐K pump either by exposing the preparation to a K‐free solution or by adding the cardioactive steroid strophanthidin increased both aiNa and twitch tension and resulted in the development of tonic tension, after‐contractions and a transient inward current (ITI). The increase of twitch tension was present at lower aiNa than that required to produce the other phenomena. The relationship between the magnitude of the twitch tension and aiNa was always non‐linear. Twitch tension increased steeply with aiNa at first but the relationship flattened off at higher aiNa and tension eventually decreased. Over the steep range, the relationship between tension and aiNa could be represented as: twitch tension = b (aiNa)y where y had a mean value of 3.2. Changing membrane potential or [Ca2+]o changed b but had little effect on y. Mn (2 mmol/l) greatly decreased twitch tension but, at least initially, had little effect on tonic tension. The steep relationship between twitch tension and aiNa was seen, irrespective of whether the Na‐K pump was inhibited either by exposure to K‐free solution or to strophanthidin and whether the relationship was measured either when aiNa was increasing or after it had reached a steady state. The steep dependence of twitch tension on aiNa observed in the present work means that manoeuvres which produce even small changes of aiNa will have significant effects on contraction.