bluuebunny/crossref_metadata_2025 huggingface dataset:
Activation by odorants of cation‐selective conductance in the olfactory receptor cell isolated from the newt.
Authors: Kurahashi T
Published: December 2014

1. Ionic selectivity of the conductance activated by n‐amyl acetate (odorant‐activated conductance) was analysed in isolated olfactory receptor cells under the whole‐cell voltage clamp condition. 2. Solitary receptor cells had a resting membrane potential of ‐44.7 +/‐ 7.0 mV (mean +/‐ S.D.; n = 70). Application of 10 mM‐n‐amyl acetate caused a depolarizing response in about 30% of the cells. Sensitivity to the odorant was maximum at around the apical dendrite. 3. Odorant induced an inward current to cells voltage clamped at their resting potential and bathed in the standard medium. The response amplitude was voltage dependent, and the polarity reversed at +2.5 +/‐ 2.2 mV (n = 6). The I‐V relation was almost linear at membrane potentials more positive than ‐20 mV, with an average slope of 3.14 +/‐ 1.59 nS (measured at 0 mV), but showed a marked outward rectification at voltages more negative than ‐30 mV. 4. Removal of external Ca2+ increased the amplitude of the odorant‐induced current and prolonged response duration, but did not cause a significant change on the reversal potential. Thus, Ca2+ affected the kinetics of the conductance, but did not seem to be a dominant charge carrier in the physiological condition. 5. Reduction of external Na+ concentration [( Na+]o) (replaced with choline) shifted the reversal potential by about 57 mV per 10‐fold change of [Na+]o. Removal of external Cl‐ (replaced with glutamate ions) did not affect the reversal potential. 6. The odorant‐activated conducting channels were permeable to all alkali metal ions. The permeability ratios were: PLi:PNa:PK:PRb:PCs = 1.25:1:0.98:0.84:0.80. 7. The present study strongly suggests that the olfactory receptor potential is generated by an increase in the membrane conductance to alkali metal ions.
Nucleotide modulation of the activity of rat heart ATP‐sensitive K+ channels in isolated membrane patches.
Authors: Lederer W J , Nichols C G
Published: December 2014

1. We have measured the ATP dependence of KATP channel activity, and the effect of various metabolites on this relationship, in inside‐out membrane patches isolated from rat ventricular myocytes. 2. The inhibition of KATP channel activity by ATP could be described as a sigmoid function of [ATP] with a Hill coefficient (HATP) of 2 and a half‐maximal inhibition at an ATP concentration (Ki, ATP) of 25 microM, in the presence of 0 mM, or 0.5 mM, total [Mg2+]. The non‐hydrolysable ATP analogue, AMP‐PNP, also inhibited the channel with Ki, AMP‐PNP = 60 microM and HAMP‐PNP = 2. 3. Acidosis caused a small, but significant, increase in Ki, ATP from 25 microM at pH 7.25 to 50 microM at pH 6.25, but phosphate and lactate were without effect (at 20 mM) on channel activity. 4. In the absence of ATP or Mg2+, ADP3‐ inhibited channel activity with Ki, ADP = 275 microM, and HADP = 1.2. Other purine and pyrimidine triphosphates, diphosphates and monophosphates also inhibited the channel with apparent order of inhibitory effectiveness ATP greater than AMP‐PNP greater than ADP greater than CTP greater than GDP = AMP = ITP. 5. In the absence of Mg2+, but in the presence of 40 microM‐ATP, channel inhibition by GTP, ITP, CTP, GDP, ADP or AMP was additive with inhibition by ATP. 6. In the presence of 0.5 mM‐Mg2+ and 40 microM‐ATP, inhibition by GTP, GMP and AMP was still additive with inhibition by ATP. The diphosphates ADP and GDP, however, paradoxically increased channel activity in the presence of ATP. This increase in channel activity appeared to result from a competitive increase in Ki, ATP, MgADP did not appear to cause any inhibition of channel activity. 7. We conclude that, in cardiac tissue, KATP channels are regulated by [ATP], and that this regulation is sensitive to other intracellular nucleotides, Mg2+, and pH, but not to phosphate or lactate. A simple, interactive two binding‐site model is consistent with the nucleotide‐dependent regulation that we observe.
Decoupling of horizontal cells in carp and turtle retinae by intracellular injection of cyclic AMP.
Authors: Miyachi E , Murakami M
Published: December 2014

1. Horizontal cells are electrically coupled through gap junctions. This is a disadvantage in elucidating the membrane properties of the cells. In order to block gap junctions, adenosine 3',5'‐cyclic monophosphate (cyclic AMP) or its analogues, dibutyryl cyclic AMP and 8‐bromo cyclic AMP, were ionophoretically injected into horizontal cells of the carp or turtle retina. 2. Before injection of the chemicals the input resistance of the cell was so low as to be unmeasurable, because the applied current leaked through gap junctions. After injection, however, the input resistance was significantly increased. 3. After the injection dye‐coupling between horizontal cells was not observed when examined by intracellular injection of Lucifer Yellow dye, supporting the idea that high concentrations of intracellular cyclic AMP block gap junctions. 4. In this situation responses to light delivered to the receptive field centre were increased in amplitude, while responses to light delivered to the receptive field surround were greatly diminished. 5. After injection horizontal cells were readily polarized by conventional intracellular current injection. The hyperpolarizing light responses in carp and turtle luminosity‐type cells (H1 cells) could be reversed by depolarizing the horizontal cells, and the reversal potentials were estimated to be about 0 mV. In addition, the resistance increase which accompanied the hyperpolarizing light responses could be detected. 6. In turtle biphasic chromaticity‐type horizontal cells (H2 cells), hyperpolarizing light responses to shorter wavelengths and depolarizing ones to longer wavelengths could be reversed by depolarizing the horizontal cells. Both responses have almost the same reversal potential at about 0 mV. The membrane resistance changes associated with light responses were also detected; the resistance increased during the hyperpolarizing response, while it decreased during the depolarizing response. These observations suggest that the ionic mechanisms of both responses are probably the same, irrespective of their polarities.
Intracellular recordings from pancreatic ganglia of the cat.
Authors: King B F , Love J A , Szurszewski J H
Published: December 2014

1. The anatomy, morphology, and electrophysiology of parasympathetic ganglia of cat pancreas were studied in vitro. 2. Pancreatic ganglia existed as an interconnected plexus of small ganglia (ten to fifty cells) lying in the interlobular connective tissue. Occasionally smaller ganglia (four to ten cells) were observed lying on or within nerve trunks. 3. Electron micrographs revealed the presence of neurones and satellite cells as well as unmyelinated axons and nerve terminals. Nerve terminals contained small clear vesicles and/or large, dense‐cored vesicles. 4. Intracellular recording of electrical activity revealed the presence of two types of ganglion cells. Type I ganglion cells exhibited resting membrane potentials that ranged from ‐40 to ‐63 mV and input resistances that ranged from 8 to 168 M omega. They responded to intracellular depolarizing current with action potentials, and received synaptic inputs which when activated caused fast and slow depolarizing responses. Type I cells were considered to be ganglionic neurones. Type II ganglion cells had higher resting membrane potentials that ranged from ‐61 to ‐83 mV, lower input resistances that ranged from 5 to 83 M omega and were electrically unexcitable. Repetitive stimulation of preganglionic nerves evoked a slow depolarization that was frequency dependent. Type II cells were considered to be satellite cells. 5. Stimulation of nerve trunks both central and peripheral to the ganglia evoked multiple, subthreshold, fast EPSPs in all type I cells tested. Fast EPSPs were blocked by the nicotinic antagonist hexamethonium. 6. Antidromic potentials were also observed following stimulation of either central or peripheral nerve trunks but never both. 7. In type I cells repetitive stimulation of both central and peripheral nerve trunks resulted in a slow, synaptically mediated depolarization which persisted during superfusion with nicotinic and muscarinic receptor antagonists. 8. Periods of low‐frequency, spontaneous fast EPSPs and action potentials were observed in all type I cells tested. 9. It was concluded that parasympathetic neurones in cat pancreatic ganglia receive convergent fast and slow synaptic inputs from central and possibly peripheral sources and may function in vivo as sites of integration. The occurrence of spontaneous synaptic potentials in pancreatic ganglia suggests the possibility of intrinsic neural control of pancreatic function.
Activation and adaptation of transducer currents in turtle hair cells.
Authors: Crawford A C , Evans M G , Fettiplace R
Published: December 2014

1. Transducer currents were recorded in turtle cochlear hair cells during mechanical stimulation of the hair bundle. The currents were measured under whole‐cell voltage clamp in isolated cells that were firmly stuck to the floor of the recording chamber. 2. Stimuli were calibrated by projecting the image of the hair bundle onto a rapidly scanned 128 photodiode array. This technique showed that, while the cell body was immobilized, the tip of the bundle would follow faithfully the motion of an attached glass probe up to frequencies of more than 1 kHz. 3. The relationship between inward transducer current and bundle displacement was sigmoidal. Maximum currents of 200‐400 pA were observed for deflections of the tip of the bundle of 0.5 microns, equivalent to rotating the bundle by about 5 deg. 4. In response to a step deflection of the bundle, the current developed with a time constant (about 0.4 ms for small stimuli) that decreased with the size of displacement. This suggests that the onset of the current was limited by the gating kinetics of the transduction channel. The onset time course was slowed about fourfold for a 20 degrees C drop in temperature. 5. For small maintained displacements, the current relaxed to about a quarter of the peak level with a time constant of 3‐5 ms. This adaptation was associated with a shift of the current‐displacement relationship in the direction of the stimulus. The rate and extent of adaptation were decreased by lowering external Ca2+. 6. Adaptation was strongly voltage sensitive, and was abolished at holding potentials positive to the reversal potential of the transducer current of about 0 mV. It was also diminished by loading cells with 10 mM of the Ca2+ chelator BAPTA. These observations suggest that adaptation may be partly controlled by influx of Ca2+ through the transducer channels. 7. Removal of adaptation produced asymmetric responses, with fast onsets but slow decays following return of the bundle to its resting position; the offset time course depended on both the magnitude and duration of the prior displacement. 8. In some experiments, hair bundles were deflected with a flexible glass fibre whose motion was monitored using a dual photodiode arrangement. Positive holding potentials abolished adaptation of the transducer currents, but had no influence on the time course of motion of the fibre. We have no evidence therefore that adaptation is caused by a mechanical reorganization within the bundle.
Further observations on the behaviour of ouabain‐insensitive sodium efflux towards proctolin in barnacle muscle fibres.
Authors: Bittar E E , Nwoga J
Published: December 2014

1. A further study has been made of the stimulatory action of proctolin on the ouabain‐insensitive Na+ efflux in single muscle fibres from the barnacle, Balanus nubilus. 2. (i) Strontium (Sr2+) behaves as a substitute for external Ca2+. In this case, however, the response to proctolin fails to decay. (ii) Injection of Sr2+ stimulates the ouabain‐insensitive Na+ efflux. This effect is mimicked by injecting Ca2+. 3. Depolarization of the fibre membrane with 30 and 100 mM‐external K+ augments the response to proctolin. 4. Pre‐injection of GTP or Gpp(NH)p (sodium 5‐guanylylimidodiphosphate) prevents the response to proctolin from completely decaying. 5. Pre‐injection of guanine nucleotides in conjunction with membrane depolarization stops the response to proctolin from decaying. 6. Measurements of Em before and during treatment with proctolin indicate a prompt but small and reversible fall in the membrane potential. 7. (i) The aequorin response of fibres pre‐treated with ouabain to proctolin is monophasic or multiphasic, and concentration dependent, the minimal effective concentration being in the nanomolar range. (ii) The duration of these signals is usually less than 5 min; this is about half the time it takes for the stimulated Na+ efflux to reach a maximum. (iii) The aequorin response to proctolin occurs quite often in fibres suspended in nominally Ca2(+)‐free artificial sea water. (iv) Sudden graded elevations in external K+ following complete decay of the aequorin response to proctolin are rapidly followed by stepwise transitory increments in light emission. (v) The aequorin response to 100 mM‐external K+ is frequently a triplet. 8. (i) Together, these results are in line with the view that the action of proctolin on the ouabain‐insensitive Na+ efflux is the result of a temporary fall in internal pCa and that its point of action is the Ca2+ channel, where a putative G protein in the presence of GTP or Gpp(NH)p is able to maintain constancy of the hormonal effect. (ii). They strengthen the argument that the barnacle muscle fibre as a preparation is especially suitable for studies of this kind.
Factors affecting aerobic recovery heat production and recovery ratio of frog sartorius.
Authors: Godfraind‐De Becker A
Published: December 2014

1. Sartorius muscles of Rana temporaria, equilibrated at 20 degrees C in Ringer solution buffered with phosphates, were stimulated isometrically for 0.2 up to 0.75 s at lengths varying from 1.03 to 1.48 times rest length, L0. The aerobic recovery heat was measured for 10.5 min after contraction. 2. The recovery heat production had a complex time course, showing a variable delay to maximum, declining thereafter. In most cases, the rate of heat production did not decrease monotonically; attention was focused on the slow exponential decay which only persisted from 1.5‐5 min after contraction. This latter part of the time course was considered as strictly aerobic and characterized by the time constant tau s. 3. Increasing the tetanus duration from 0.2 to 0.75 s increased initial heat Qi and recovery heat Qr in proportion, so that the recovery ratio R (Qr/Qi) did not change; it was equal to 1.29 +/‐ 0.03 (S.E.M.; n = 44) for muscles at about L0. The kinetics of heat production were modified with longer tetani; in particular, tau s was increased from 2.2 to 5.2 min. 4. When muscles were stretched beyond L0, as long as there was no increase of the resting heat rate (stretch response or ‘Feng effect’), recovery heat production had a similar evolution to that in muscles at about L0; R was constant and equal to 1.21 +/‐ 0.03 (n = 46). 5. When muscles were sufficiently stretched to develop a stretch response, R increased proportionally to the stretch response. The effect seemed independent of the contractile machinery, as it vanished concomitantly with the stretch response, while force and Qi remained unchanged for the length considered. The kinetics were also modified‐‐the delay to maximum was no longer detected and tau s most likely increased. 6. Substitution of 60% of the NaCl of the Ringer solution by NaI (mol/mol) produced a significant increase of R, mainly due to the increase of Qr. 7. The results show that neither the time course nor the amount of aerobic recovery heat Qr are strictly determined by the amount of initial heat Qi. The hypothesis is discussed that Qr might include a variable fraction due to processes which are not directly implicated in the actin‐myosin interactions, possibly those involving the cytosolic Ca2+ concentration and the rate of resting metabolism.
Synovial capillary distribution in relation to altered pressure and permeability in knees of anaesthetized rabbits.
Authors: Levick J R , McDonald J N
Published: December 2014

1. The hydraulic conductance of the synovial lining of the rabbit knee increases greatly at intra‐articular pressures (IAP) above 9 cmH2O. A structural cause was sought by fixing synovium in situ at less than or equal to 5 cmH2O IAP (ten animals) or 25 cmH2O IAP (five animals) and examining histological sections morphometrically. 2. The synovial lining was found to be a highly deformable sheet of very vascular connective tissue, with 47 x 10(3)‐73 X 10(3) capillaries per cm2 section. 150‐260 cm2 endothelial surface per cm3 tissue and a vascular volume of 2.4‐5.7%. 3. The thickness of the lining averaged 14‐19 microns at low IAP and was reduced at high IAP; in suprapatellar synovium, where changes were most marked, thickness fell by 24‐47%. The loose subsynovial space expanded. 4. The average distance separating capillary near‐edges from the joint cavity approximately halved from 3.75 and 7.47 microns at low IAP (harmonic and arithmetical means respectively) to 1.82 and 3.35 microns at high IAP. Capillaries remained patent and their number density did not change significantly at high IAP. 5. It is concluded that a reduction in the extravascular path length for fluid exchange contributes to the increase synovial conductance at high IAP, but the path length changes were not sufficient to account fully for the conductance changes.
The steady‐state and rebreathing methods compared during morphine administration in humans.
Authors: Bourke D L , Warley A
Published: December 2014

1. We examined the quantitative and qualitative differences between the steady‐state and rebreathing methods of determining CO2‐response curves before and after each of two doses of intravenous morphine (0.07 and 0.14 mg kg‐1) in four healthy male humans. 2. During each study session steady‐state and rebreathing CO2‐response curves were determined as an ordered pair (separated by 15 min). Carbon dioxide‐response curves were determined for control, after 0.07 mg kg‐1 morphine, and after a total of 0.21 mg kg‐1 morphine. Each subject was studied on a second occasion when the order of the CO2‐response pairing was reversed. 3. The results are discussed and related to a model that may account for the differences based on the step increase in CO2 at the onset of rebreathing, the rate of rise of CO2 during rebreathing and the time constant for the central chemoreflex. 4. Our empirical conclusion is that morphine causes a parallel right shift of the steady‐state CO2‐response curve and causes a non‐specific decrease in the slope of the rebreathing CO2‐response curve. We suggest that the parallel shift of the steady‐state CO2‐response curve is specific to drugs acting on opioid receptors.
The effects of lesions on autogenetic inhibition in the decerebrate cat.
Authors: Iles J F , Jack J J , Kullmann D M , Roberts R C
Published: December 2014

1. The effects of spinal and brain lesions on autogenetic inhibition from contraction receptors were studied in the decerebrate cat. Inhibitory feedback gain was estimated by measuring the effect of tension perturbations on reflex contractions of the soleus muscle. Tendon vibration was used to clamp the firing rate of primary spindle afferents, to prevent spindle unloading from disfacilitating the reflex contraction. In addition, secondary spindle afferents could be selectively excited by stimulating fusimotor fibres during muscle vibration. 2. Following an acute contralateral or bilateral dorsal transection of the spinal cord at L3, the vibration reflex tension fell by between 50 and 74% in three decerebrate animals. This was accompanied by a variable increase in inhibitory feedback, ranging between 180 and 360%. 3. In two animals, selective stimulation of fusimotor fibres supplying soleus muscle was without effect in the presence of muscle vibration both before and after the spinal lesion. In the third animal, a small and variable reduction in tension could be obtained only after the lesion, implying that an inhibitory pathway from homonymous secondary spindle afferents to alpha‐motoneurones was released. 4. In a separate series of experiments, contralateral cerebral lesions were made 2‐12 months prior to the acute inhibitory feedback measurement. Inhibitory feedback gain was increased, on average twofold in decerebrate animals with chronic cerebral lesions, when compared to control decerebrate animals. 5. Selective stimulation of fusimotor fibres to excite spindle secondary afferents was uniformly without effect in decerebrate animals with chronic cerebral lesions. In one animal spinal transection had only a minor effect on extensor tone and on inhibitory feedback gain, in contrast to the control decerebrate cats. 6. The implications of these findings are discussed in relation to the use of animals with spinal and supraspinal lesions as models of spasticity.
Electrical and mechanical activity of rabbit prostate smooth muscles in response to nerve stimulation.
Authors: Seki N , Suzuki H
Published: December 2014

1. The rabbit prostate contains two smooth muscle bundles: one forms the capsule of the prostate and the other runs longitudinally in the outermost layer of the prostate. The former was contracted by exogenously applied noradrenaline (NA) and the latter by acetylcholine (ACh), and were therefore identified as NA‐sensitive and ACh‐sensitive muscles, respectively. 2. In the NA‐sensitive muscle, exogenously applied NA, but not ACh, depolarized the smooth muscle membrane, and this action of NA was antagonized by prazosin but not by yohimbine. The membrane of the ACh‐sensitive muscle was depolarized by ACh and hyperpolarized by NA. The former was antagonized by atropine and the latter by phentolamine. 3. In both bundles, electrical stimulation of intramural nerves elicited a biphasic junction potential: an excitatory junction potential (EJP) followed by an inhibitory junction potential (IJP). In the NA‐sensitive muscle, the EJP was resistant to alpha‐adrenoreceptor antagonists and sensitive to guanethidine, while the IJP was sensitive to apamin and resistant to alpha‐ or beta‐adrenoreceptor antagonists or guanethidine. The EJP and IJP elicited in the ACh‐sensitive muscle were inhibited by atropine and apamin, respectively, but both potentials were resistant to alpha‐ or beta‐adrenoreceptor antagonists or guanethidine. 4. Transmural nerve stimulation elicited a twitch contraction in both muscle bundles, and the contractions in the NA‐sensitive muscle were partially inhibited by prazosin and blocked completely by guanethidine. The contraction of the ACh‐sensitive muscle was blocked by atropine. Contractions produced by exogenously applied NA and ACh were antagonized by prazosin and atropine, respectively. 5. It is concluded that in the rabbit prostate there are two smooth muscles bundles which are respectively contracted with associated depolarization by NA or ACh. The NA‐sensitive muscle receives adrenergic excitatory and non‐adrenergic non‐cholinergic inhibitory innervation. The ACh‐sensitive muscle receives cholinergic excitatory and non‐adrenergic non‐cholinergic inhibitory innervation.
The initiation of calcium release following muscarinic stimulation in rat lacrimal glands.
Authors: Marty A , Tan Y P
Published: December 2014

1. Acinar cells were isolated from rat lacrimal glands, and the Ca2+ release response of these cells was studied using two experimental approaches. In one approach, changes in Ca2+ concentration, Cai2+, were monitored by measuring Ca2(+)‐dependent Cl‐ currents using tight‐seal whole‐cell recording. Alternatively, such changes were measured as a fluorescence signal in cells loaded with Fura‐2. 2. Following bath application of ACh (0.5 microM), the cell current recorded at ‐60 mV was unchanged for ca 0.8 s, then rose in a biphasic manner. The initial phase of the current rise ('hump') took different appearances depending on the cell studied, and it sometimes stood out from the main part of the response as a partially isolated transient. 3. In cells which had been loaded with Fura‐2, Cai2+ was found to rise abruptly following a silent period. The delay was larger if ACh (0.2‐0.5 microM) was applied in a depolarizing isotonic K+ saline than if it was applied in the normal saline. In addition, the maximum of the Cai2+ response was reduced with depolarizing stimulating solutions. This indicates that membrane potential modulates the Cai2+ response. 4. Responses to 5 microM‐ACh, a saturating agonist concentration, were almost identical in K+ saline and in normal saline. 5. If the cell potential was hyperpolarized, the delay of the ACh‐induced current became shorter. 6. Breaking into an acinar cell with a pipette containing an elevated Ca2+ concentration (0.1‐1 mM) led to a transient activation of Ca2(+)‐induced currents during the first seconds of whole‐cell recording. These transients were obtained more reliably if the transition to the whole‐cell mode was achieved by applying a sharp pulse of potential ('zapping') rather than by applying suction to the pipette compartment. At ‐60 mV, the transients elicited with the former method by 0.5 mM‐Ca2+ had a time‐to‐peak near 0.6s and an amplitude varying between 10 and 600 pA. With 0.1 mM‐Ca2+, similar transients were also observed, but a number of cells failed to respond. Calcium‐induced transients were blocked if cells were previously loaded with 50 microM‐Ruthenium Red. 7. Performing the same experiments with inositol trisphosphate (InsP3, 20 microM) in the pipette solutions also led to early transient Ca2(+)‐induced currents. Amplitudes, times‐to‐peak and 20‐80% transition times were similar for 0.5 mM‐Ca2+ and 20 microM‐InsP3 stimulations.(ABSTRACT TRUNCATED AT 400 WORDS)
Gating of the afferent volley of the monosynaptic stretch reflex during movement in man.
Authors: Meunier S , Pierrot‐Deseilligny E
Published: December 2014

1. The time course of the changes in presynaptic inhibition of Ia fibres to soleus motoneurones has been investigated during a voluntary ramp‐and‐hold plantar flexion. 2. Monosynaptic Ia facilitation of the soleus H reflex was evoked by stimulation of the inferior soleus (homonymous) and femoral (heteronymous) nerves. Changes in presynaptic inhibition of the Ia fibres mediating the conditioning volleys were inferred from changes in the amount of reflex facilitation evoked by such constant conditioning stimulations. 3. At the beginning of the voluntary contraction, both homonymous and heteronymous Ia facilitations were markedly increased with respect to their rest values. Then, they dropped abruptly in the middle of the ramp, whatever the ramp duration. 4. The stronger the contraction at the end of the ramp, the larger the initial increase in reflex facilitation. 5. These results indicate that presynaptic inhibition of Ia terminals on motoneurones of the contracting muscle is decreased during the first half of the ramp and then returns to its rest level. It is argued that these changes in presynaptic inhibition are centrally programmed, and it is suggested that, under certain conditions, presynaptic inhibition can be modulated according to both the strength and timing of the contraction. 6. The functional role of the resulting changes in the gain of the stretch reflex is discussed.
Putative cerebral cortical involvement in the ventilatory response to inhaled CO2 in conscious man.
Authors: Murphy K , Mier A , Adams L , Guz A
Published: December 2014

1. The response of the diaphragm to both transcranial magnetic stimulation and electrical phrenic nerve stimulation was studied in thirteen normal subjects under conditions of either a ‘reflex’ drive to ventilation with inhaled CO2 or during volitional ventilation of similar magnitude. 2. The induced compound action potential in the diaphragm was recorded using an oesophageal electrode, and in some cases transdiaphragmatic pressure was recorded using oesophageal and gastric balloon catheters. 3. The response of the diaphragm to transcranial magnetic stimulation was invariably facilitated with volitional inspiration; there was either minimal or no response at functional residual capacity. 4. Facilitation with inspiration was also seen during a ‘reflex’ drive to ventilation with inhaled CO2 in the presumed absence of any volitional contribution to ventilation. A similar degree of facilitation was seen with voluntary ventilation of similar magnitude and pattern. 5. If the facilitation is predominantly a cortical phenomenon, then these results imply that there is a behavioural component in the previously supposed purely ‘reflex’ drive to ventilation with inhaled CO2. We also discuss the interpretation of these results if some of the facilitation occurs at the phrenic motoneurone.
The uptake of radiolabelled precursors of mucus glycoconjugates by secretory tissues in the feline trachea.
Authors: Davies J R , Corbishley C M , Richardson P S
Published: December 2014

1. We gave one of three radiolabelled precursors of mucus glycoconjugates ([3H]proline, [3H]glucose and [35S]sulphate) into the tracheas of anaesthetized cats for 3 h. In other cats [35S]sulphate was given by intravenous injection. 2. After a further 2 h, tracheas were removed and fixed. Serial actions were cut and alternate sections were stained with Haematoxylin and Eosin or prepared as unstained autoradiographs. Points on submucosal gland and surface epithelium were chosen with a grid on photomicrographs of the stained sections. Absorbance, which is proportional to autoradiographic grain density, was estimated on corresponding points on unstained autoradiographs by flying‐spot microdensitometry. 3. With [3H]proline as precursor, the grain densities were greater over surface epithelium than over submucosal gland. With [3H]glucose, grain densities were greater over the surface epithelium in three cases, equal in one and greater over submucosal gland in the last. [35S]Sulphate, given either into the tracheal segment or intravenously, yielded grain densities that were greater over the submucosal glands than over surface epithelium. 4. The areas of submucosal gland the surface epithelium were estimated by point counting and the total content of radioactivity in the two structures estimated by multiplying mean absorbance by area. Ratios of the total radiolabel in surface epithelium to that in submucosal gland were consistently high when [3H]proline was the precursor and low with [35S]sulphate, given by either route. [3H]Glucose gave intermediate ratios. 5. Secretions washed from the trachea were subjected to gel‐exclusion chromatography. Washings from tracheas labelled with [3H]proline contained some molecules eluting in the void volume of a Sepharose CL‐4B column (suggesting a relative molecular mass of greater than 10(6) Da), but more of the radiolabel eluted in three peaks in the partially included volume. Density gradient ultracentrifugation of the void volume material gave radiolabelled peaks at densities of approximately 1.60 and 1.50 g ml‐1, consistent with glycosylated proteins, as well as less dense material (less than 1.30 g ml‐1), probably proteins with little or no glycosylation. 6. We discuss the justification of using these radiolabelled precursors to give relatively selective labelling of secretory products from submucosal gland and surface epithelium.
Agonist‐evoked changes in cytosolic pH and calcium concentration in human platelets: studies in physiological bicarbonate.
Authors: Sage S O , Jobson T M , Rink T J
Published: December 2014

1. Cytosolic pH (pHi) and calcium concentration ([Ca2+]i) have been investigated in the presence and absence of physiological HCO3‐ in human platelets co‐loaded with the fluorescent indicators BCECF and Fura‐2. Basal pHi and changes evoked by butyrate, thrombin, platelet activating factor (PAF), ADP and phorbol ester were investigated, as were the effects of removing external Na+. 2. In the presence of physiological HCO3‐ and CO2, basal pHi was 7.02 +/‐ 0.04 compared with 7.15 +/‐ 0.05 in the absence of HCO3‐. Estimated cytosolic buffering power was reduced from 35.6 +/‐ 3.0 to 14.5 +/‐ 0.4 mM/pH unit by the omission of HCO3‐. 3. Thrombin evoked an immediate acidification of 0.03 +/‐ 0.01 pH units in the presence of HCO3‐ and 0.07 +/‐ 0.01 pH units in its absence. The acidifications were followed by a slow alkalinization. The final pHi was 0.10 +/‐ 0.01 units above basal in the presence of HCO3‐ and 0.08 +/‐ 0.02 units above basal in the absence of HCO3‐. The initial acidification was significantly greater in the absence of HCO3‐. The subsequent increase in pHi was similar in the presence and absence of this ion, but the calculated loss of proton equivalents was greater in the presence of HCO3‐. 4. Replacement of extracellular Na+ with N‐methyl‐D‐glucamine resulted in a fall in basal pHi and abolished recovery from thrombin‐evoked acidification in both the presence and absence of HCO3‐. 5. In the presence of HCO3‐, PAF and ADP evoked an intracellular acidification similar to that caused by thrombin. However, with PAF and ADP, the subsequent recovery in pHi was slow and did not rise above basal levels. Phorbol dibutyrate, an activator of protein kinase C, evoked a similar elevation in pHi of 0.04 +/‐ 0.01 units over 3 min in the presence and absence of HCO3‐. 6. Stopped‐flow fluorimetric measurements were made of both BCECF and Fura‐2 fluorescence in the presence of HCO3‐. In the presence and absence of external Ca2+, thrombin‐evoked rises in [Ca2+]i peaked before any cytoplasmic alkalinization occurred. ADP evoked rapid elevations in [Ca2+]i, but caused no alkalinization.(ABSTRACT TRUNCATED AT 400 WORDS)
Two transient outward currents in histamine neurones of the rat hypothalamus in vitro.
Authors: Greene R W , Haas H L , Reiner P B
Published: December 2014

1. The transient outward current exhibited by the histamine neurones of the tuberomammillary nucleus was studied using the single‐electrode voltage clamp technique in an in vitro rat hypothalamic slice preparation. 2. The transient outward current exhibited steady‐state inactivation at the resting potential. Inactivation was removed by priming hyperpolarization with a V1/2 of ‐85 +/‐ 1.2 mV, while the V1/2 for activation was ‐60.3 +/‐ 2.1 mV. 3. The decay of the transient outward current was best fitted by two exponentials with time constants of 104 +/‐ 36 and 568 +/‐ 128 ms. These two components were provisionally termed IA,f and IA,s for the fast and slowly decaying currents, respectively. 4. Removal of inactivation was time dependent; inactivation was fully removed by hyperpolarizing pulses to ‐110 mV of 200 ms or greater duration. Removal of inactivation of IA,f was rapid, becoming complete with hyperpolarizing pre‐pulses of 50 ms or greater, while removal of inactivation of IA,s was not complete until hyperpolarizing pre‐pulses were 200 ms in duration. 5. The fast decaying current IA,f was selectively blocked by 1 mM‐4‐aminopyridine. Tetraethylammonium chloride (10 mM) had no effect on either IA,f or IA,s. 6. The inactivation curves for IA,s, determined both by using the values obtained from the amplitude of the computed slower exponential function as well as that of the current remaining in 1 mM 4‐aminopyridine, were negative to those of IA,f. Similarly derived activation curves for IA,s were positive to those of IA,f. 7. Superfusion with a nominal 0 Ca2+ medium containing 10 mM‐Mg2+ did not reduce the maximal transient outward current. 8. The reversal potential of IA,s with 2.5 mM‐K+ in the medium was ‐95 +/‐ 3 mV; the reversal potential of IA,f was at least 15 mV negative to that of IA,s. 9. It is concluded that histaminergic tuberomammillary neurones possess at least two transient outward currents which can be distinguished on the basis of their rates of decay, 4‐aminopyridine sensitivity, voltage dependence and reversal potentials.
Differential block of sodium and calcium channels by chlorpromazine in mouse neuroblastoma cells.
Authors: Ogata N , Yoshii M , Narahashi T
Published: December 2014

1. The effects of chlorpromazine on the voltage‐activated sodium and type I (transient) calcium channels were studied in cultured mouse neuroblastoma cells (N1E‐115) using the whole‐cell patch‐clamp technique. 2. Chlorpromazine (2‐10 x 10(‐6) M) blocked both the sodium channel current and the calcium channel current as carried by Ba2+ in a reversible and dose‐dependent manner. 3. The block was not associated with any change in the time course of the activation and inactivation of the sodium and calcium channel currents. 4. The dose‐response relationships for the block of these channels measured with a holding potential of ‐120 mV indicated a 1:1 binding stoichiometry with apparent dissociation constants of 2.5 +/‐ 10(‐6) M and 1.5 +/‐ 10(‐5) M for the sodium and calcium channels, respectively. 5. The block was dependent on the holding potential for both channel currents. The apparent dissociation constant for the sodium channel was decreased to 0.65 +/‐ 10(‐6) M when the membrane was held at ‐80 mV. The apparent dissociation constant for the calcium channel was decreased to 3.2 +/‐ 10(‐6) M when the membrane was held at ‐60 mV. 6. The steady‐state inactivation curve for the sodium channel was shifted by 12.4 +/‐ 1.8 mV to more negative potentials by exposure to 1 x 10(‐6) M‐chlorpromazine. The inactivation curve for the calcium channel was also shifted by 15.4 +/‐ 3.2 mV to more negative potentials by exposure to 1 x 10(‐5) M‐chlorpromazine. These results indicate a greater affinity of chlorpromazine for the inactivated state of the channels than for the resting state. 7. Chlorpromazine caused a marked use‐dependent block of the sodium channel current, as demonstrated by a cumulative increase of the block during a train of depolarizing pulses. The use‐dependent block was observed even with an interpulse interval as long as 2 s. 8. On the other hand, the block of the calcium channel current did not notably accumulate during a train of depolarizing pulses even when extremely prolonged (1 s) pulses were applied at a very short interpulse interval (200 ms). 9. The marked use dependence of the sodium channel block was due to a very slow repriming of the drug‐bound sodium channels from inactivation, whereas the lack of use dependence of the calcium channel block was due to a rapid repriming of the drug‐bound calcium channels.(ABSTRACT TRUNCATED AT 400 WORDS)
GABAA receptor function is regulated by phosphorylation in acutely dissociated guinea‐pig hippocampal neurones.
Authors: Chen Q X , Stelzer A , Kay A R , Wong R K
Published: December 2014

1. Current mediated by GABAA receptors was examined in pyramidal cells acutely dissociated from the hippocampus of mature guinea‐pigs. Current responses were measured using whole‐cell voltage‐clamp recordings. An internal perfusion technique was used to change the intracellular contents during recording. 2. Application of GABA (100‐300 microM) by short duration pressure pulses produced outward current responses at a holding potential of ‐10 mV. When recordings were made with intracellular solutions which did not contain Mg‐ATP, GABA responses progressively decreased to less than 10% of their initial values after 10 min. This ‘run‐down’ of the GABA response could not be accounted for by desensitization since the rate of run‐down was not dependent upon agonist application. 3. The run‐down of the GABAA response was reversed when Mg2+ (4 mM) and ATP (2 mM) were introduced into the intracellular perfusate. In addition to the presence of Mg‐ATP, buffering of Ca2+ in the intracellular solution to low levels (approximately 10(‐8) M) was also necessary to stabilize the GABAA response. 4. The role of a phosphorylation process in regulating the GABAA receptor was tested. After the GABA response stabilized, introduction of alkaline phosphatase (100 micrograms/ml) to the intracellular perfusate caused a complete run‐down of the GABA response. 5. Stable GABA responses were obtained when ATP was replaced by ATP‐gamma‐S (adenosine 5'‐O‐(thiotriphosphate), an analogue of ATP that donates a thiophosphate group resulting in a product that is more resistant to hydrolysis. Following such treatment GABA responses declined more slowly after the introduction of intracellular alkaline phosphatase. 6. Run‐down of GABA responses accelerated when intracellular Ca2+ concentration ([Ca2+]i) was elevated to about 5 x 10(‐4) M. The run‐down caused by elevated [Ca2+]i could be stopped and reversed by reducing [Ca2+]i to about 10(‐8) M. 7. The introduction of ATP‐gamma‐S to the intracellular medium retarded the run‐down of GABA responses caused by elevation of [Ca2+]i. 8. N‐(6‐Aminohexyl)‐5‐chloro‐1‐naphthalenesulphonamide (W‐7), a calmodulin inhibitor, reduced the rate of run‐down induced by elevated [Ca2+]i. 9. These results suggest that the function of the GABAA receptor is maintained by phosphorylation of the receptor or some closely associated regulatory molecule. Elevation of [Ca2+]i destabilizes the function of the GABAA receptor, probably by activating a Ca2+/calmodulin‐dependent phosphatase.
Multiple components of both transient and sustained barium currents in a rat dorsal root ganglion cell line.
Authors: Boland L M , Dingledine R
Published: December 2014

1. Currents through voltage‐activated Ca2+ channels in rat dorsal root ganglion (DRG) x mouse neuroblastoma hybrid (F‐11) cells were studied using the whole‐cell patch clamp technique with 30 mM‐Ba2+ as charge carrier. Two components of the inward Ba2+ current were distinguished on the basis of voltage dependence and time course. Each component could be further subdivided based on pharmacology. 2. A transient inward current activated at test potentials positive to ‐40 mV, peaked within 20 ms and then decayed during a 200 ms depolarization. The peak amplitude of the transient current occurred between ‐10 and +10 mV. With a 300 ms conditioning pulse, half‐inactivation of the transient current occurred at ‐30 mV. A sustained inward current activated at test potentials positive to ‐30 mV and reached a maximum at +20 to +30 mV. The sustained current showed little voltage‐dependent inactivation over 200 ms. The amplitudes of both the transient and sustained currents were increased by perfusing with Ba2+ instead of Ca2+. 3. Most F‐11 cells had both the transient and sustained Ba2+ currents although the relative amount of the two currents varied with culture conditions. The transient current was more prominent in cells fed with a ‘growth’ medium (15‐20% serum) whereas the sustained current was increased in cells fed with a ‘differentiation’ medium (1% serum plus growth factors). F‐11 cells can be used to study transient current in relative isolation from sustained Ca2+ current under certain culture conditions. The neuroblastoma parent of the F‐11 cell line, N18TG‐2 cells, exhibited little or no voltage‐dependent Ba2+ current. 4. Brief application of omega‐conotoxin fraction GVIA (10 microM) produced a long‐lasting block of 81% of the sustained current and 27% of the transient current. 5. The transient and sustained Ba2+ currents in F‐11 cells were reversibly blocked by brief exposure to Cd2+ or Ni2+. Block of the sustained current was evident with 100 nM‐Cd2+ whereas the threshold concentration for Ni2+ block was 1 microM. Cd2+ and Ni2+ were equipotent blockers of the transient current. Dose‐response curves for Cd2+ and Ni2+ block of both sustained and transient currents had shallow slopes suggesting that the block was more complex than a simple bimolecular interaction between blocker and one blocking site. Dose‐response curves were fitted by a model that included two binding sites for each divalent blocker.(ABSTRACT TRUNCATED AT 400 WORDS)
Vasodilatation of arterioles by acetylcholine released from single neurones in the guinea‐pig submucosal plexus.
Authors: Neild T O , Shen K Z , Surprenant A
Published: December 2014

The nervous control of arterioles in the guinea‐pig submucosal plexus was studied. Outside diameters of arterioles were recorded using a video‐monitoring system. Changes in arteriolar diameter in response to electrical stimulation of single neurones or ganglia in the plexus were measured. 2. When the arteriole was pre‐constricted with the prostaglandin analogue U46619 or with phenylephrine, electrical stimulation (2‐20 Hz, 10 s) of a ganglion dilated the blood vessel. This vasodilatation was abolished by tetrodotoxin or by cutting the fine nerve strands running between the ganglion and the arteriole. 3. The vasodilatations caused by ganglionic stimulation were blocked by the muscarinic antagonists atropine, pirenzepine, (11[[2‐[(diethylamino)methyl]‐1‐piperidinyl]acetyl]‐5,11‐dihydro‐6H‐ pyrido[2,3‐b][1,4]benzodiazepine‐6‐)‐one (AFDX‐116), 4‐diphenylacetoxy‐N‐methyl‐piperidine methiodide (4‐DAMP) and hexahydrosilodifenidol (HSDF). IC50 values for the inhibition of nerve‐evoked vasodilatation by pirenzepine, AFDX‐116 and HSDF were 500 nM, 4 microM and 25 nM respectively. Physostigmine (1 microM) increased the dilatation by 90%. 4. Muscarine dilated all submucosal arterioles; the concentration causing half‐maximum effects was 200 nM. Muscarinic vasodilatations were inhibited by pirenzepine, AFDX‐116, and HSDF in a competitive manner; dissociation equilibrium constants determined by Schild analyses were 125 nM, 1.3 microM and 4 nM respectively. 5. Gossypol, an irreversible inhibitor of the production of endothelium‐derived relaxing factor (EDRF), did not reduce the vasodilatation produced by either ganglionic stimulation or muscarine in submucosal arterioles. 6. Intracellular recordings were made from submucosal neurones and action potentials were elicited by depolarizing current pulses (10 ms duration, 10 Hz/10 s). In seven neurones vasodilatation was associated with intracellularly evoked action potentials; this vasodilatation was blocked by pirenzepine. Cell bodies of reidentified vasodilator neurones were subsequently shown to contain immunoreactive choline acetyltransferase. 7. These results show that cholinergic neurones in the submucosal plexus project to submucosal arterioles and that they release acetylcholine onto muscarinic receptors to produce vasodilatation. The muscarinic receptor activated by nerve‐released acetylcholine is the M3 subtype and its location appears to be on the vascular smooth muscle rather than the endothelium.