bluuebunny/crossref_metadata_2025 huggingface dataset:
Electrical membrane responses to secretagogues in parietal cells of the rat gastric mucosa in culture.
Authors: Okada Y , Ueda S
Published: December 2014

Fragments of the gastric fundus of 6‐8‐day‐old rats were maintained in tissue culture. From the explant, adhered to a plastic substrate, epithelial cells migrated and developed to form a monolayer colony. Histological and histochemical studies as well as indirect immunofluorescence studies using anti‐parietal cell antibodies testified to the presence of parietal cells in the monolayer during the first week. These parietal cells were distinguished by their vesicular cytoplasmic structures using phase‐contrast or differential interference‐contrast microscopy. Acridine Orange, an optical probe of H+ accumulation, was taken up preferentially by these parietal cells, exhibiting orange fluorescence within the cells on the third day of culture, in response to stimulation with gastrin, histamine and carbachol. The resting potential of these cultured parietal cells was about ‐20 mV. On day 2‐4 of culture, the cell membrane became hyperpolarized (up to ‐30 to ‐40 mV) in response to gastrin, carbachol or histamine in the presence of isobutylmethyl‐xanthine (IMX). During hyperpolarization, the membrane resistances decreased significantly. The amplitude and the polarity of secretagogue‐induced responses were found to be dependent on the extracellular concentration of K+ (but not Na+ and Cl‐). The carbachol‐induced responses were inhibited by atropine but not curare. The responses induced by histamine plus IMX were blocked by cimetidine but not pyrilamine. Neither atropine nor cimetidine affected the gastrin‐evoked responses. It is concluded that rat parietal cells have separate receptors for acetylcholine (muscarinic), gastrin and histamine (H2), and that an increase in the membrane permeability to K+ is closely associated with the responses of these receptors under these in vitro conditions.
Sodium‐ and adenosine‐triphosphate‐dependent calcium movements in membrane vesicles prepared from dog erythrocytes.
Authors: Ortiz O E , Sjodin R A
Published: December 2014

Inside‐out vesicles from the membranes of dog erythrocytes were obtained by the method of Lew & Seymour (1982) for study of Ca movements. In the absence of ATP, 45Ca accumulation by the vesicles was inhibited by external Na and stimulated by internal Na. The presence of either MgCl2, quinidine sulphate, or LaCl3 in the incubation medium inhibited 45Ca accumulation in the absence of ATP. The release of 45Ca from 45Ca‐loaded vesicles was specifically promoted by Na+ in the absence as well as in the presence of ATP. The accumulation of 45Ca by vesicles was stimulated by ATP and the effect of ATP was entirely dependent on the presence of Mg. The Mg‐ and ATP‐dependent 45Ca accumulation was stimulated by the presence of either K or Na in the medium, was hyperbolically activated by increasing the Ca2+ concentration in the medium, was stimulated by calmodulin and inhibited by orthovanadate (10(‐4) M) or LaCl3 (10(‐3) M). The data demonstrate the presence of two mechanisms for controlling Ca movements in inside‐out vesicles from dog erythrocyte membranes, a Na‐dependent one similar to the Na‐Ca exchange described for squid axons and cardiac muscle and a Ca pump utilizing ATP with characteristics similar to those described for human erythrocytes and squid axons.
Control of the repetitive discharge of rat CA 1 pyramidal neurones in vitro.
Authors: Madison D V , Nicoll R A
Published: December 2014

Experiments using intracellular recording techniques were performed on rat hippocampal neurones in vitro, to study the discharge properties of these cells. When CA 1 pyramidal cells were excited by injecting long depolarizing current pulses (approximately 600‐800 ms), they responded with an initial rapid action potential discharge which slowed, or accommodated, and then stopped after 200‐300 ms. The train of action potentials was followed by a hyperpolarization which was due primarily to calcium‐activated potassium conductance (GK(Ca]. The amplitude of this hyperpolarization increased with an increasing number of action potentials in the initial discharge. Blocking the calcium‐activated potassium conductance, by injecting EGTA into the cell, by bathing the cell in cadmium, a calcium channel blocker, or by bathing the cell in calcium‐free medium, reduced the after‐hyperpolarization (a.h.p.) and accommodation such that the frequency of action potential discharge increased and the duration of this discharge was prolonged. Blocking the calcium‐activated potassium conductance had a greater effect on discharge frequency later in the action potential train, as late interspike intervals were shortened more than early ones by the application of cadmium or of calcium‐free medium. This was presumably because the calcium‐activated potassium conductance was more developed later in the train. Accommodation was not completely abolished in the absence of calcium and presence of cadmium, suggesting that other factors, in addition to calcium‐activated potassium conductance, contributed to this process. This remaining accommodation was reduced by low doses of carbachol, suggesting that the M‐current also plays a role in accommodation. We conclude that accommodation of the action potential discharge of hippocampal pyramidal cells may be regulated by at least two potassium currents: the calcium‐activated potassium current and the M‐current. Both of these currents are turned on during excitation of the neurone and act in an inhibitory manner on that neurone to limit further action potential discharge.
Non‐dopaminergic neurones of the reticular part of substantia nigra can gate static fusimotor action onto flexors in cat.
Authors: Schwarz M , Sontag K H , Wand P
Published: December 2014

The effect on the fusimotor system of electrical stimulation of the reticular part of the substantia nigra or of the injection of the gamma‐aminobutyric acid (GABA)‐antagonist picrotoxin into this structure was studied in spindle receptors of pretibial flexors in cats anaesthetized with ketamine. Afferent activity of muscle spindle primary endings was recorded before and during these two forms of intranigral stimulation. Dynamic spindle sensitivity was assessed during both small‐ (100 microns) and large‐amplitude (2 mm) sinusoidal stretching of the receptor‐bearing muscles. From changes in spindle sensitivity after nigral electrical stimulation (eleven out of fourteen primary endings) or intranigral injection of picrotoxin (fifty‐one out of sixty‐seven primary endings) it is deduced that functional activation of neurones of the reticular part of substantia nigra, in this preparation, removed a normally present tonic static fusimotor action from the primary sensory endings of pretibial flexor muscle spindles. This effect, induced by picrotoxin (2 micrograms in 1 microliter), was reversed by a subsequent intranigral injection of the GABA‐agonist muscimol (0.4 microgram in 1 microliters), but remained unchanged after subsequent intracaudate injections of haloperidol (12.5 micrograms in 5 microliters) or apomorphine (5 micrograms in 5 microliters). It is concluded that the C.N.S. can gate static fusimotor action onto flexor muscle spindle primary endings through non‐dopaminergic output neurones of the reticular part of substantia nigra.
Arsenazo III calcium transients and latency relaxation in frog skeletal muscle fibres at different sarcomere lengths.
Authors: Close R I , Lännergren J I
Published: December 2014

Single, intact, frog muscle fibres were injected electrophoretically with a Ca2+‐sensitive metallochromic dye, Arsenazo III, to a local concentration of 1.2‐1.5 mmol/l. The intracellular concentration of free Mg2+, estimated photometrically in the presence of approximately millimolar Arsenazo III, was 3‐4 mmol/l in fibres at rest. Ca2+‐related changes in dye absorbance were characterized in vitro using 1 mM‐Arsenazo III in solutions approximating the intracellular ionic environment. Isometric twitch contractions and related changes in light transmittance of dye‐injected regions of fibre were recorded at 2.4 and 3.0 micron sarcomere lengths, at 15 degrees C. A method was developed for separating Ca2+ transients from larger, movement‐related optical changes recorded as compound signals during fibre contraction. Decreases in twitch amplitude by about one‐third following dye injection, together with the in vitro characteristics of the dye, suggested that millimolar intracellular Arsenazo III acted as a major Ca2+ buffer and inhibited the activation of contractile filaments. The onset of both the Ca2+ transient and latency relaxation occurred at virtually the same time in the twitch response and neither of those transition times was altered significantly with changes in sarcomere length from 2.4 to 3.0 micron. The amount of activation Ca2+ released in dye‐injected regions of fibres following a single stimulus was about 0.3 mmol/l at 2.4 micron sarcomere length. The rate of rise and the amplitude of the Ca2+ transient were reversibly decreased with increase in sarcomere length from 2.4 to 3.0 micron. That finding is reviewed in relation to other evidence indicating length dependence of the intracellular release and distribution of activation Ca2+ up to 3.9 micron sarcomere length.
Initiation of migrating myoelectric complex in sheep by duodenal acidification and hyperosmolarity: role of vagus nerves.
Authors: Gregory P C , Rayner D V , Wenham G
Published: December 2014

Gastrointestinal motility was studied in conscious sheep by X‐radiography and by electromyography from chronically implanted electrodes before and after total thoracic vagotomy. Duodenal infusion of 0.5‐3 mmol HCl (0.035‐0.1 M‐HCl) induced premature duodenal regular spiking activity (r.s.a.) within 1‐7 min in fifteen of seventeen sheep studied when infused at 20 min after a natural r.s.a. There was no correlation between abomasal pH and any phase of the migrating myoelectric complex (m.m.c.). Duodenal alkalinization by infusion of 0.3 M‐Tris buffer (pH 10.2) or 0.1 M‐NaHCO3 had no influence on the occurrence of the m.m.c. Duodenal infusion of 20‐50 ml 0.5 M‐NaCl induced a premature duodenal r.s.a. within 1‐5 min in seven of eight sheep. Vagotomy did not prevent the initiation or migration of the m.m.c., but reduced the rate of propagation of the r.s.a. from 40.5 +/‐ 7.2 (mean +/‐ S.E. of mean) to 16.7 +/‐ 0.1 cm/min in the duodenum, from 27.3 +/‐ 4.1 to 16.6 +/‐ 0.8 cm/min in the jejunum, and from 21.4 +/‐ 1.1 to 13.7 +/‐ 0.7 cm/min in the proximal ileum. Initially the frequency of r.s.a. increased, especially in the duodenum where they recurred at an interval of 98.4 +/‐ 6.8 min before vagotomy; and at 23.4 +/‐ 1.8 min in the first 24 h after vagotomy; the interval had lengthened to 86.7 +/‐ 5.2 min 2‐3 weeks after vagotomy. Premature duodenal r.s.a. was not induced by duodenal infusion of HCl in five, or by duodenal infusion of hyperosmolar NaCl in three chronically vagotomized sheep. It is concluded that the vagus nerves contribute to the regulation of the frequency and propagation of the m.m.c. in sheep; duodenal acidification is not essential nor is it the normal stimulus for initiation of r.s.a., but duodenal infusion of HCl or hyperosmolar NaCl can initiate a premature duodenal r.s.a. via the vagus nerves.
The effects of caffeine on tension development and intracellular calcium transients in rat ventricular muscle.
Authors: Konishi M , Kurihara S , Sakai T
Published: December 2014

The effects of caffeine on tension and intracellular [Ca2+] were investigated in rat ventricular muscle using the Ca2+‐sensitive photoprotein, aequorin. Contracture was induced by rapid application of 0.5‐10 mM‐caffeine solution at 20 degrees C. In normal Tyrode solution at 8 degrees C, or in Na+‐deficient solution in which Na+ was isotonically replaced by sucrose, peak tension of caffeine contracture was potentiated and relaxation was prolonged. Caffeine contracture could not be induced immediately after a prior contracture. Repriming time was 10 min in Tyrode solution, and was much shorter in Na+‐deficient solution or in high‐K+ solution containing 105.9 mM‐K+. Caffeine prolonged the plateau of action potential dose dependently. At low temperature, prolongation of the plateau phase by caffeine was more marked. Twitch tension showed a triphasic change after application of caffeine; peak tension transiently increased in a potentiating phase (P phase), and then decreased below control level in an inhibitory phase (I phase) followed by gradual recovery in a recovery phase (R phase). The effects of caffeine on the Ca2+ transients during a twitch were also complex, depending on time after application and dose of caffeine. In low caffeine concentration (below 0.5 mM) the peak of the Ca2+ transient was potentiated in the I phase, although the peak tension was suppressed. At high concentration (above 3 mM) the peaks of both the Ca2+ transient and twitch tension were suppressed. In every concentration of caffeine tested (0.1‐5 mM), time to the Ca2+ transient and twitch tension peaks was prolonged, and the falling phases of both were delayed. Caffeine might release Ca2+ from intracellular store(s) and enhance the slow inward current. The Ca2+ transient obtained in this study clearly indicate that the prolonged time to peak tension in the presence of caffeine is due to the slow rise of intracellular [Ca2+] and prolonged time to peak of the Ca2+ transient. It is also quite possible that caffeine modulates the Ca2+ sensitivity of a contractile system in dose‐ and time‐dependent manners.
A transient sodium‐hydrogen exchange system induced by catecholamines in erythrocytes of rainbow trout, Salmo gairdneri.
Authors: Baroin A , Garcia-Romeu F , Lamarre T , Motais R
Published: December 2014

The addition of isoprenaline to an isotonic suspension of red blood cells of rainbow trout induces an amiloride‐sensitive Na+ transport which is independent of Cl‐ and insensitive to 4,4'‐diisothiocyano‐2,2'‐stilbene disulphonic acid (DIDS) and furosemide. Na+ uptake is accompanied by amiloride‐sensitive H+ release. The H+ efflux is dependent upon the external Na+ concentration, the K0.5 value for Na+ being 16 mM. In the presence of DIDS, when the coupled NaCl entry (NaCl co‐transport) induced by catecholamine is blocked, the results provide evidence for a linked movement of Na+ and H+, with a stoicheiometry of 1:1. Exchange of H+ for Na+ induces osmotic swelling of the cells which is due to the replacement of a bound proton by an osmotically active Na+ cation. In the absence of DIDS when the bulk of the Na+ uptake is the result of a coupled entry of Na+ and Cl‐, H+ extrusion still occurs and the magnitude of acid excretion is identical to that found in DIDS‐treated cells. This suggests that Na+‐H+ exchange remains active. Addition of isoprenaline first stimulates the Na+‐H+ exchange but only transiently. This is followed by a more permanent stimulation of the NaCl co‐transport.
Gustatory trophic action of arterial chemosensory neurones in the cat.
Authors: Dinger B , Fidone S J , Stensaas F J
Published: December 2014

The proximal stump of the carotid sinus nerve (c.s.n.) branch of the IXth cranial nerve was surgically cross‐anastomosed to the distal stump of the lingual nerve (l.n.) branch of the same cranial nerve to test the ability of regenerating c.s.n. axons to reinnervate and induce taste buds on cat circumvallate papillae. The l.n. branch of the IXth nerve, which normally innervates circumvallate papillae, was directed away from the tongue by suturing it to the distal stump of the c.s.n. Animals with normal and re‐anastomosed (cut, then resutured) l.n. served as control preparations. 2‐19 months following cross‐anastomosis, circumvallate papillae contained 59.8 +/‐ 9.1% (mean +/‐ S.E. of mean) of the normal incidence of taste buds, indicating that axons possessing specific gustatory trophic properties had reinnervated the epithelium. Further experimentation was concerned with confirming the identity of the putative foreign axons which reinnervated the taste buds. Radiolabelled axoplasmic transport and autoradiography showed that 89 +/‐ 4.4% of the taste buds in cross‐anastomosed, and 97 +/‐ 2.1% in normal preparations were innervated by axons from the principal sensory ganglion (petrosal) of the IXth nerve (P greater than 0.05). Whole‐nerve recordings in cross‐anastomosed preparations demonstrated gustatory activity in the transposed c.s.n., which was similar to responses present in normal and re‐anastomosed l.n. preparations. The largest response was evoked by 4 M‐NaCl, followed by 1 M‐NH4Cl and 0.02 M‐quinine HCl. Sucrose (0.5 M) elicited insignificant nerve activity. In normal animals, gustatory or mechanical stimulation did not alter cardiopulmonary function, but in bilaterally cross‐anastomosed preparations, gustatory stimuli produced respiratory excitation while mechanical stimulation resulted in reductions in blood pressure and respiratory inhibition. These data suggest that following cross‐anastomosis, arterial chemosensory axons innervate taste buds and barosensory axons innervate tongue mechanoreceptors (which normally evoke cardiopulmonary reflexes from the carotid body and carotid sinus, respectively). We conclude that arterial chemosensory neurones share with gustatory chemosensory neurones a trophic function essential for the development and maintenance of taste buds.
Physiological role of apical potassium ion channels in frog skin.
Authors: Van Driessche W
Published: December 2014

The K+ permeability of the apical membrane of frog skin (Rana temporaria) was analysed by recording the short‐circuit current and its fluctuations in the presence of a mucosa‐to‐serosa‐oriented K+ concentration gradient. Loading of the animals with KCl resulted in an augmentation of the Ba2+‐blockade component of the short‐circuit current and the plateau value of the K+‐dependent relaxation noise. Poisoning of active transport and exposing both sides of the epithelium to KCl Ringer solution caused an increase of the K+ current and its fluctuations recorded after restoring the inward‐oriented K+ gradient. Serosal quinidine (5 X 10(‐4) M), which is thought to increase intracellular Ca2+ activity, depressed the K+ current and the relaxation noise. This effect was completely reversible. Removal of Na+ from the serosal solution, which is known to result in an elevation of intracellular Ca2+ by abolishing the driving force for the Na+/Ca2+ exchanger, also reduced the K+ current and the Lorentzian plateau. Both parameters returned to their control values after restoring the Na+ gradient across the basolateral membranes. It is concluded from these experiments that the apical K+ permeability is controlled by factors which depend on the intracellular K+ and Ca2+ concentration and that the apical K+ channels may constitute a pathway for K+ secretion.
Beta‐adrenergic receptor mechanisms in rat parotid glands: activation by nerve stimulation and 3‐isobutyl‐1‐methylxanthine.
Authors: Fuller C M , Gallacher D V
Published: December 2014

The technique of electrical field stimulation (e.f.s.) was employed in conjunction with selective pharmacological antagonists to specifically investigate the role of endogenous neurotransmitter(s) in the activation of beta‐adrenergic receptor mechanisms in isolated parotid gland segments of the rat. The field‐stimulus‐induced amylase release due to beta‐adrenergic receptor activation was characterized as that persisting in the presence of atropine (10(‐5) M) and phentolamine (10(‐5) M) and susceptible to blockade by propranolol (5 X 10(‐6) M), i.e. combined beta 1‐ and beta 2‐receptor blockade. The selective beta 1‐receptor antagonist metoprolol (10(‐5) M) was as effective as propranolol in blocking the beta‐mediated enzyme release. The selective beta 2‐receptor antagonist, H35/25 (10(‐5) M) did not significantly affect the field‐stimulus‐induced amylase release. In the absence of any phosphodiesterase inhibitor the levels of cyclic AMP in the tissues were close to the limit of detection. Field stimulation was however associated with a fourfold increase in cyclic AMP. By comparison isoprenaline (10(‐5) M) gave rise to a tenfold increase in cyclic AMP. The changes in cyclic AMP metabolism, in response to both field stimulation and isoprenaline, were greatly enhanced in the presence of the phosphodiesterase inhibitor 3‐isobutyl‐1‐methylxanthine (IBMX). The field‐stimulus‐induced increase in cyclic AMP was abolished by the beta 1‐adrenergic receptor antagonist, metoprolol, but persisted in the presence of the beta 2‐adrenergic antagonist, H35/25. IBMX was found to have a potent direct effect on amylase release. IBMX (10(‐3) M) also gave rise to a tenfold increase in cyclic AMP. IBMX is then as effective as 10(‐5) M‐isoprenaline in stimulating both enzyme secretion and cyclic AMP metabolism. The secretory response to IBMX was unaffected by beta‐adrenergic blockade by propranolol, was independent of extracellular calcium and did not give rise to 86Rb+ efflux. Importantly, isoprenaline (10(‐5) M) failed to evoke any significant increase in amylase release if introduced during sustained superfusion of IBMX, yet it is in such protocols that the greatest changes in cyclic AMP metabolism are seen. The study clearly demonstrates that the beta‐adrenergic‐receptor‐regulated amylase release in response to nerve stimulation is mediated predominantly, if not exclusively, by the beta 1‐receptor subtype.(ABSTRACT TRUNCATED AT 400 WORDS)
Muscle but not cutaneous C‐afferent input produces prolonged increases in the excitability of the flexion reflex in the rat.
Authors: Wall P D , Woolf C J
Published: December 2014

Stimulation of cutaneous afferent fibres in the sural nerve and muscle afferent fibres in the gastrocnemius‐soleus nerve at a strength that excites C axons produces a delayed and long‐lasting burst of activity in posterior biceps femoris/semitendinosus flexor motoneurones. Following a 20 s stimulation at 1 Hz to the sural nerve the flexor motoneurones continue to fire for 20 s while a similar stimulus to gastrocnemius‐soleus nerve results in an after‐discharge lasting three times longer. Using stimuli to the sural and gastrocnemius‐soleus nerves as conditioning stimuli (20 s, 1 Hz) changes in the excitability of the flexor reflex were measured by recording the discharge evoked by a test sural nerve stimulus or by a standard pinch to the ipsilateral and contralateral toes. Prior to any conditioning stimulus the flexor reflex remained stable for prolonged periods. Conditioning stimuli at strengths that activated large myelinated afferent fibres only, or large and small myelinated afferent fibres, failed to produce more than a very transient alteration in the reflex excitability. Conditioning stimuli at C‐fibre strength to the sural nerve produced a marked increase in the excitability of the reflex for 10 min. C‐fibre strength gastrocnemius‐soleus nerve conditioning stimuli resulted in a similar increase in excitability followed by a second phase of facilitation peaking at 20‐30 min and lasting for up to 90 min. The afferent barrage initiated by cutting the sural and gastrocnemius‐soleus nerves resulted in similar patterns of reflex excitability increases with the muscle nerve resulting in a more prolonged effect than the cutaneous nerve. The results show that a brief C‐afferent fibre input into the spinal cord can produce a prolonged increase in the excitability of the flexion reflex and that muscle C‐afferent fibres evoke longer‐lasting changes than cutaneous C fibres. The differences in the time course of the post‐conditioning effects may be related to the well‐described differences in the sensory consequences of injury to skin versus deep tissue.
Effects of acetylcholine and short‐chain fatty acids on acinar cells of the exocrine pancreas in sheep.
Authors: Katoh K , Tsuda T
Published: December 2014

In order to investigate the actions of acetylcholine and short‐chain fatty acids on acinar cells of the exocrine pancreas of sheep, measurements of amylase release and 45Ca efflux from superfused segments, as well as changes in membrane potential, input resistance and equilibrium potential in the acinar cells, were carried out in vitro. The application of acetylcholine or short‐chain fatty acids caused a dose‐dependent increase in amylase release from the superfused tissue segments. The amylase release evoked by 10(‐3) M‐short‐chain fatty acids containing 2‐8 carbon atoms increased with increasing carbon number, up to 5 (i.e. it was maximum with iso‐valerate, which has 5 carbon atoms). The amylase release stimulated by acetylcholine (5.5 X 10(‐6) M) or caprylate (10(‐3) was accompanied by an increase in 45Ca efflux, and was significantly reduced by the removal of extracellular Ca2+. The stimulating effect of caprylate (10(‐3) M) on amylase secretion was also observed in superfused segments of the guinea‐pig, but not in those of the mouse, rabbit or hamster. The resting membrane potential and input resistance of acinar cells of sheep pancreas were ‐31.1 +/‐ 1.6 mV and 2.7 +/‐ 0.7 M omega (means +/‐ S.E. of means), respectively. The application of acetylcholine or short‐chain fatty acids always depolarized the cell membrane and reduced the input resistance. The effect of short‐chain fatty acids was observed in the presence of atropine (1.4 X 10(‐6) M). The equilibrium potentials for acetylcholine and butyrate were ‐15.0 +/‐ 0.8 and ‐16.0 +/‐ 1.1 mV, respectively. It is concluded that the cellular secretory process evoked by acetylcholine is qualitatively similar to that of short‐chain fatty acids, and that Ca2+ ions might be an important mediator for these secretagogues in the acinar cells of the pancreas of sheep.
Long‐term facilitation of peptidergic transmission by catecholamines in guinea‐pig inferior mesenteric ganglia.
Authors: Dun N J , Jiang Z G , Mo N
Published: December 2014

Intracellular recordings were obtained from neurones of isolated guinea‐pig inferior mesenteric ganglia. Repetitive stimulation (10‐20 Hz for 1‐2 s) of the hypogastric nerves evoked, in addition to the fast excitatory post‐synaptic potential (e.p.s.p.), a non‐cholinergic e.p.s.p. the mediator of which has previously been suggested to be substance P or a related peptide. When applied to the ganglia in the concentrations of 1‐100 microM for 3‐5 min, adrenaline, isoprenaline and noradrenaline produced an initial, short‐lasting depression which was followed by a marked augmentation of the non‐cholinergic e.p.s.p. lasting from minutes to over hours. Employed in comparable concentrations dopamine caused a slight depression that was not followed by a detectable increase of the non‐cholinergic e.p.s.p. The catecholamine‐induced depression and subsequent enhancement of the non‐cholinergic e.p.s.p. was prevented by alpha‐adrenergic antagonists (dihydroergotamine and phenoxybenzamine, 1‐10 microM) and beta‐adrenergic antagonists (propranolol and dichlorisoprenaline, 5‐10 microM), respectively. The membrane depolarization induced by the putative transmitter substance P (1 microM) was augmented by isoprenaline; the enhancement which could be blocked by beta‐antagonists was not preceded by a depression. Application of dibutyryl cyclic AMP (10 microM‐1 mM) by either superfusion or intracellular ionophoresis mimicked the enhancing effect of catecholamines. It is concluded that catecholamines, with the noticeable exception of dopamine, exerted a biphasic effect on the non‐cholinergic e.p.s.p. of the inferior mesenteric ganglion cells: an initial depression that was mediated by alpha‐adrenergic receptors and probably reflected a presynaptic inhibitory effect of catecholamines and, on the other hand, an enduring facilitation mediated by beta‐adrenergic receptors which appeared to be linked to activation of post‐ganglionic cyclic AMP.
On the basis of delayed depolarization and its role in repetitive firing of Rohon‐Beard neurones in Xenopus tadpoles.
Authors: Spitzer N C
Published: December 2014

A delayed depolarization following the impulse can be recorded intracellularly from mature Rohon‐Beard neurones in the spinal cord of Xenopus tadpoles, in response both to brief intracellularly injected current pulses and to antidromic stimulation. Evidence is presented suggesting that this delayed depolarization is unlikely to be due to the action of a chemical synapse, activation of a voltage‐dependent conductance in the cell body, increased extracellular potassium, or electrotonic coupling. Hyperpolarization of the cell body during antidromic stimulation eliminates the action potential normally generated there, and reveals an impulse arising at some distance along a neurite. When an action potential is produced in the cell body, its repolarizing phase sculpts a delayed depolarization from this impulse in the neurite. The depolarization is enhanced by pressure applied to the neurites near the cell body, presumably by reducing the distal spread of current, and yields multiple action potentials. Although long current pulses usually produce only a single spike, small quantities of La3+ enhance the size of the depolarization and cause repetitive firing. The relation of impulse frequency to injected current shows a non‐linearity consistent with the summation of the delayed depolarization and the depolarization by the injected current. The non‐linearity is eliminated upon removal of delayed depolarization by hyperpolarizing current pulses injected after each impulse. The enhancement of the depolarization by La3+ is not the only cause of repetitive firing; La3+ also produces an effective reduction in conductance for outward currents. This depolarization may play a role in the normal firing behaviour of Rohon‐Beard neurones; when repetitive firing results naturally in response to long current pulses the delayed depolarization is observed to be large.
Release of adenosine, inosine and hypoxanthine from rabbit non‐myelinated nerve fibres at rest and during activity.
Authors: Maire J C , Medilanski J , Straub R W
Published: December 2014

The composition of the efflux from desheathed rabbit vagus nerve, loaded with radioactivity by incubation in [3H]adenosine, was studied at rest and during electrical activity and after application of inhibitors of ecto‐enzymes and modifications of intermediary metabolism. In addition, the degradation of externally applied ATP and adenosine was examined. [3H]ATP applied to the incubation medium was degraded to ADP, AMP, adenosine and inosine. The hydrolysis to nucleosides was inhibited by alpha, beta‐methylene ADP; the appearance of AMP and nucleosides was slowed by beta, gamma‐methylene ATP. Deamination of [3H]adenosine was blocked by 2‐deoxycoformycin. The effluent from resting and stimulated preparations showed the presence of large amounts of inosine and hypoxanthine, smaller amounts of adenosine and adenine and traces of nucleotides. The composition of the effluent was not significantly altered by addition of alpha, beta‐methylene ADP; beta, gamma‐methylene ATP or 2‐deoxycoformycin. Application of glucose‐free solutions caused a large release of adenosine instead of inosine and hypoxanthine and a small increase in resting and stimulated efflux of 3H. Addition of 2‐deoxyglucose produced a large increase in resting efflux and increased liberation of adenosine. Cyanide, 2,4‐dinitrophenol, arsenate or salicylate increased the resting efflux of adenosine, inosine and hypoxanthine, and the effect of activity. It is concluded that electrical activity leads to release of adenosine, inosine and hypoxanthine, in various proportions depending on metabolic state, and that there is practically no liberation of nucleotides from nerve axons.
Effect of alpha‐dihydro‐grayanotoxin‐II on the electrical activity of the rabbit sino‐atrial node.
Authors: Nakao M , Seyama I
Published: December 2014

The effect of alpha‐dihydro‐grayanotoxin‐II (GTX) on the electrical activity of rabbit sino‐atrial (s.a.) node cells was studied by the two‐micro‐electrode voltage‐clamp technique. GTX, at concentrations between 3 and 10 microM, suppressed the spontaneous beating of the s.a. node. On subsequent application of 1 microM‐tetrodotoxin (TTX), the membrane repolarized and spontaneous beating was restored, whereas the maximum rate of rise and the frequency of the action potential were reduced slightly. The additional application of adrenaline (0.55 microM) (in the presence of GTX plus TTX) completely restored the normal electrical activity of the s.a. node cells. Voltage‐clamp experiments revealed that GTX, in this concentration range, reduced the maximum conductance of slow inward current by 15%, and did not affect the outward current system. The steady‐state inactivation curve for the slow inward current was not shifted along the membrane potential axis, whereas that for the fast inward Na current was shifted in the direction of hyperpolarization. In addition, GTX generated a time‐independent current (IGTX) which could be eliminated by the application of TTX. The current‐voltage relation for IGTX was linear and crossed the voltage axis at +4.0 +/‐ 2.2 mV (n = 4). The application of GTX in a concentration range above 30 microM abolished all gated inward and outward currents of the s.a. node. The results suggest that the GTX‐induced sinus arrest is mainly due to an increase in the membrane permeability to Na ions and that this increase in permeability is due to conversion of the normal fast Na channel into a modified one, which lacks an inactivation process.
Innervation of the muscularis mucosae of canine proximal colon.
Authors: Angel F , Go V L , Szurszewski J H
Published: December 2014

The innervation of the muscularis mucosae of the canine large intestine was studied in vitro using superfusion and radioimmunological techniques. In the majority of preparations, electrical field stimulation (10 V, 200 microseconds, 10 Hz) elicited a biphasic neurogenic response which consisted of a contraction followed, after cessation of the stimulus, by relaxation. Electrical field stimulation released VIP‐, substance P‐ and bombesin‐like immunoreactivity. Release of these peptides and the biphasic response to nerve stimulation were blocked by tetrodotoxin and a 'calcium‐free' solution. Several observations suggest that neuronally released substance P (or a closely related peptide) mediated the contraction by a direct action on the muscle. The contraction caused by substance P was tetrodotoxin insensitive. Desensitization to substance P abolished the excitatory response to nerve stimulation. The contraction elicited by nerve stimulation was blocked by substance P antiserum. Several observations suggest that bombesin or a closely related peptide caused contraction of the muscle by releasing substance P from intramural neurones. Bombesin caused an increase in substance P‐like immunoreactivity in the superfusate which was blocked by tetrodotoxin, as was the contraction; substance P antibodies blocked the contractile response to bombesin. In addition, while the excitatory response to electrical nerve stimulation was blocked by substance P antiserum, there was still an increase in bombesin‐like immunoreactivity in the superfusate. The data also suggest that VIP or a closely related peptide might have mediated the relaxation by a direct action on the muscle. The inhibitory response to nerve stimulation was mimicked by VIP and abolished by VIP antiserum.
Chloride current induced by injection of calcium into Xenopus oocytes.
Authors: Miledi R , Parker I
Published: December 2014

Membrane currents of Xenopus oocytes were studied with the membrane under voltage clamp. Intracellular injection of the calcium‐chelating agent EGTA reduced, or abolished, the transient outward chloride current normally activated by membrane depolarization. Intracellular injection of calcium ions evoked large membrane currents, which inverted direction close to the chloride equilibrium potential. Injections of strontium, or barium, were less effective than calcium, while magnesium was ineffective. Large chloride currents could be evoked by calcium injections in oocytes which showed only small or no transient outward currents. The current activated by calcium injection increased with increasing depolarization up to high (ca. +60 mV) positive potentials, even though the transient outward current was suppressed by strong depolarization. The results indicate that the transient outward current depends upon entry of calcium through voltage‐gated calcium ion channels and show that the oocyte membrane contains numerous chloride channels which are activated by intracellular calcium. Only a few of these chloride channels are activated by depolarization.
Arsenazo III used as a calcium ion indicator in mouse oocytes.
Authors: Hunter M J
Published: December 2014

The Ca2+‐indicator dye Arsenazo III has been injected into mouse oocytes by intracellular ionophoresis, and the optical absorbance of the injected dye measured using a double‐beam microspectrophotometer. The absorbance spectrum shows a broad peak between 565 and 580 nm, with a maximum of 0.020‐0.095 in different oocytes. From these values, intracellular dye concentrations were estimated to be 0.1‐0.5 mM, although estimated concentrations up to 1.0 mM have been used in later experiments where the dye absorbance was measured only at selected wave‐lengths. The transport number for intracellular ionophoresis of the dye has been estimated. The mean value was 0.009 in experiments where the injection current was allowed to cross the cell membrane, but was only 0.0015 when the injection current was balanced by a current of opposite polarity through a second electrode barrel. In oocytes exposed to the mitochondrial uncoupler DNP (2,4‐dinitrophenol), the shape of the Arsenazo III absorbance spectrum was found to change. The resulting difference spectra show the peaks at about 610 and 660 nm which are characteristic features of the Arsenazo III Ca2+‐difference spectrum. These absorbance changes were not completely reversible on removal of DNP. Changes in the optical absorbance of Arsenazo III have been observed in oocytes injected with Ca2+ by intracellular ionophoresis. Estimated increases of intracellular free [Ca2+], which were of the order of 1 microM and decayed within 30 s, are discussed in relation to the Ca2+‐buffering and Ca2+‐elimination properties of the oocyte.
Development of gamma‐aminobutyric acid mediated inhibition of X cells of the cat lateral geniculate nucleus.
Authors: Berardi N , Morrone M C
Published: December 2014

We studied the development of gamma‐aminobutyric (GABA) mediated inhibitory processes of the lateral geniculate nucleus cells in kittens of various age groups, by measuring the effect of ionophoretic application of GABA and bicuculline on cell response to sinusoidal gratings. In young kittens (less than 30 days) we found very few Y cells only X cells and weakly responsive cells which fitted neither X nor Y classifications ('immature' cells). As with adult cells, GABA inhibits the visual response of young kitten cells. Simultaneous application of bicuculline restored responsiveness. The mean GABA current to silence cell response in young kittens was significantly higher than that obtained in adult cats. Application of bicuculline alone had little effect on young kitten X‐cell responsiveness, either on the average discharge or on the amplitude of modulation to stimulation by sinusoidal gratings. For older kittens (40‐45 days), bicuculline increased X‐cell responsiveness, and the increase in responsiveness was dependent on stimulus contrast and spatial frequency. However, the increased responsiveness was less than that for adults. At 100 days the changes of slope of X‐cell contrast‐response curves during bicuculline application were similar to those observed for adult X cells. We conclude that, although GABA receptors may be present at 30 days, the GABA mediated inhibitory system does not begin to function until about 45 days and does not mature fully until about 100 days.
A mechanical study of regulation in the striated adductor muscle of the scallop.
Authors: Simmons R M , Szent-Györgyi A G
Published: December 2014

Chemically skinned fibre bundles were prepared from the striated adductor muscle of the sea scallop, Placopecten magellanicus. The relation between tension and calcium concentration was determined in activating solutions containing 5 mM‐MgATP, ionic strength 0.2, pH 7.1 at 20 degrees C. The isometric tension rose from zero to its maximum value between pCa 6.0 and 5.2. The steepness of the relation cannot be accounted for in terms of the binding of calcium to the two known sites on myosin and suggests that there must be an additional, co‐operative mechanism. The regulatory light chain content of the fibre bundles was determined by urea gel electrophoresis and was found to be approximately 2 light chains per myosin molecule. The regulatory light chains were removed completely by treatment with EDTA at 25‐30 degrees C. Fibre bundles then showed a total loss of control over contraction; a high tension was generated whether or not calcium was present in the bathing solution. Complete removal of the regulatory light chains did not greatly affect the tension generated or the stiffness in the rigor state. Control of contraction could be restored completely by the addition of regulatory light chains from scallop muscle. Treatment with EDTA at 0‐12 degrees C resulted in the removal of 0.76‐2.0 regulatory light chains per myosin molecule. Fibre bundles for which removal was less than complete were partially sensitive to calcium, i.e. tension was higher in the presence of calcium than in its absence. The results indicate that the normal mechanism of tension generation in scallop muscle is mediated primarily through myosin and not thin filament control. This finding is consistent with previous studies of the ATPase activity of myofibrils from scallop muscle.
The effect of autonomic agonists and nerve stimulation on protein secretion from the rat submandibular gland.
Authors: Jones C J , Wilson S M
Published: December 2014

Saliva samples were obtained from the cannulated duct of the rat submandibular gland following intravenous administration of the autonomic agonists, acetyl beta‐methyl choline (methacholine), phenylephrine and isoprenaline and by stimulating sympathetically at 5 and 20 Hz. Phenylephrine and isoprenaline produced a saliva approximately 50‐fold richer in protein than that induced by methacholine. Protease activity was greatest in saliva induced by phenylephrine. Methacholine, phenylephrine and isoprenaline caused the secretion of markedly different protein populations, determined by separation on SDS‐polyacrylamide gradient gels followed by staining with Coomassie Brilliant Blue R250. Many species in the molecular weight range 10000‐40000 were secreted in response to phenylephrine and isoprenaline. Methacholine produced a saliva poor in protein and the resulting electrophoretic banding patterns were faint. Staining with periodic acid‐Schiff's reagent showed that the largest protein species (mol. wt. 150000) secreted in response to isoprenaline was glycosylated and that small quantities of a similar protein were present in saliva produced by methacholine and phenylephrine but were not visualized by staining with Coomassie Brilliant Blue R250. Phosphorylated proteins of molecular weights 30000‐35000 and 38000‐45000 were detected in saliva produced by isoprenaline but not in saliva produced by methacholine or phenylephrine. Stimulation of the cervical sympathetic trunk at 5 Hz yielded a population of salivary proteins, many of which behaved similarly on electrophoresis to proteins secreted in response to isoprenaline. At 20 Hz a different population was secreted, resembling that present in phenylephrine‐induced saliva.
A voltage‐clamp analysis of membrane currents in solitary bipolar cells dissociated from Carassius auratus.
Authors: Kaneko A , Tachibana M
Published: December 2014

Membrane properties of solitary bipolar cells, mechanically dissociated from the enzyme‐treated goldfish retina, were studied under current‐ and voltage‐clamp conditions with 'giga‐seal' suction pipettes (pipette solution 138 mM‐K). The resting potential of solitary bipolar cells was about ‐30 mV. They responded to depolarizing current pulses with sustained depolarization, and to hyperpolarizing current pulses with an initial hyperpolarizing transient followed by a sag to a less hyperpolarized level. The current‐voltage relationship determined under voltage‐clamp conditions showed strong outward and inward rectification. The membrane currents consisted of four components; Ca current (ICa), voltage‐ and Ca‐dependent K currents (IK(V) and IK(Ca), respectively), and an inward current activated by membrane hyperpolarization (Ih). ICa was activated by membrane depolarization beyond ‐40 mV, was maximum at +10 mV and became smaller with further depolarization. No polarity reversal was seen. ICa was enhanced by equimolar replacement of Ca with Ba, and was blocked by 4 mM‐Co. IK(Ca) was observed by membrane depolarization beyond ‐10 mV, was maximum at about +40 mV, and became smaller with further depolarization. This current was suppressed by 4 mM‐Co, 1.6 mM‐Ba, 35 mM‐TEA or 30 microM‐quinine. IK(V) was activated by membrane depolarization beyond ‐60 mV, and had slower kinetics that ICa or IK(Ca). The reversal potential of the tail current was close to the K equilibrium potential (EK), suggesting that this current is carried purely by K ions. IK(V) was inactivated slowly and nearly completely by sustained depolarization. IK(V) was blocked by 35 mM‐TEA. Ih was activated by membrane hyperpolarization (less than ‐60 mV). The current showed a time‐dependent increase. It was also dependent on the membrane potential, but not on the driving force of K ions. This current seems to be carried by a mixture of Na and K ions, since (1) in low Na solution, Ih became small in amplitude, and (2) the reversal potential of the tail current was between the Na equilibrium potential (ENa) and EK X Ih was blocked by 10 mM‐Cs, but was resistant to 0.2 mM‐Ba. The resting potential and voltage responses of solitary bipolar cells are discussed in reference to the characteristics of each membrane conductance isolated in the present study.
Abdominal vascular responses to changes in carbon dioxide tension in the cephalic circulation of anaesthetized dogs.
Authors: Ford R , Hainsworth R , Rankin A J , Soladoye A O
Published: December 2014

Dogs were anaesthetized with chloralose, the regions of both carotid sinuses were vascularly isolated and perfused with arterial blood and both cervical vagosympathetic trunks were cut above the nodose ganglia. The cephalic circulation was perfused through the brachiocephalic and left subclavian arteries with blood which was equilibrated with various levels of CO2. The abdomen was vascularly isolated, perfused through the aorta at constant flow and drained through the inferior vena cava at constant pressure. Changes in vascular resistance were determined from changes in abdominal aortic perfusion pressure and changes in capacitance from the integral of the changes in venous outflow. An increase in PCO2 in the cephalic perfusate resulted in an increase in abdominal vascular resistance and a decrease in capacitance. However, when carotid sinus pressure was high, the response of resistance to an increase in cephalic PCO2 was abolished and that of capacitance was significantly reduced. The reflex responses of both vascular resistance and capacitance to a change in carotid sinus pressure were enhanced when the cephalic PCO2 was raised. However, the effect on the reflex capacitance response from stimulation of baroreceptors was obtained only when PCO2 was changed below 5 kPa whereas the effect on resistance occurred at higher values of PCO2. The interaction between the effects of changes in cephalic PCO2 and the carotid sinus reflex and the differential effect on resistance and capacitance vessels have been explained in terms of the known difference in the sensitivities of these vessels to sympathetic nerve activity.