bluuebunny/crossref_metadata_2025 huggingface dataset:
Inhibition of the calcium pump by high cytosolic Ca2+ in intact human red blood cells.
Authors: Pereira A C , Samellas D , Tiffert T , Lew V L
Published: December 2014

1. The inhibitory effect of high intracellular calcium on the saturated Ca2+ efflux through the Ca2+ pump (Vmax) was investigated in intact human red cells. Cells were loaded with Ca2+ by exposure to the calcium ionophore A23187, at different external Ca2+ concentrations ([Ca2+]o). Ca2+ extrusion by the pump was followed after either ionophore removal or Co2+ addition. 2. fifty per cent inhibition of Vmax was obtained with total intracellular calcium ([CaT]i) of approximately 3 mmol/l cells. For any given initial Ca2+ load, Vmax showed no tendency to increase as [CaT]i was progressively reduced during Ca2+ efflux. This suggests that the pump Vmax was determined by the magnitude of the initial [Ca2+]i. 3. To estimate [Ca2+]i from [CaT]i in Co(2+)‐loaded cells, the possible competition between Co2+ and Ca2+ for the known cytoplasmic Ca2+ buffers (alpha‐buffers) was investigated first. Comparison between Ca2+ efflux after either Co2+ exposure or ionophore wash‐out showed that the efflux patterns were essentially identical, down to the lowest measurable [CaT]i. This indicates that Co2+ does not compete with Ca2+ for the alpha‐buffers. Hence, since [Ca2+]i = alpha [CaT]i, and alpha approximately 0.15‐0.35, the initial [Ca2+]i load for 50% Vmax inhibition was between 0.4 and 1.1 mM. 4. Ancillary new findings demonstrated that, unlike the situation with alpha‐buffers, Co2+ displaced Ca2+ from the cell‐incorporated calcium chelator benz‐2, and that benz‐2 incorporation had no effect on Co(2+)‐exposed Ca2+ pump desaturation. This validates the use of benz‐2 to study Ca2+ pump kinetics in intact cells.
Inter‐individual variability in the dynamics of natural accommodation in humans: relation to age and refractive errors.
Authors: Schaeffel F , Wilhelm H , Zrenner E
Published: December 2014

1. To study the relationship between accommodation under natural viewing conditions, age and refractive errors, we have measured time courses of accommodation in thirty‐nine human subjects aged 5‐49 years using a newly developed technique. The technique is based on infrared photoretinoscopy and involves fully automated on‐line image processing of digitized video images of the eyes with a sampling rate of 5.3 Hz. 2. The distance between the subject and the video camera was about 1.3 m. Head movements of the subject required little restriction because the eyes were automatically tracked in the video image by the computer program. All subjects were tested under binocular viewing conditions. 3. Both refraction of the right eye and pupil diameter were measured with a precision of 0.2‐0.4 dioptres (D) and 0.1 mm, respectively, and were plotted on‐line. The data were subsequently automatically analysed. 4. Automated infrared photoretinoscopy proved to be very convenient and easy to handle in both children and adults. 5. The maximal speed of accommodation for a target at a distance of 5 D declined in the subjects with age (from up to 21.7 D s‐1 for accommodation and 32.7 D s‐1 for subsequent accommodation to a distant target ('near to far accommodation') in children down to 2‐18 D s‐1 in adults). There was a striking inter‐individual variability in the maximum possible speed of accommodation and near to far accommodation. 6. Speed of accommodation and of near to far accommodation was correlated for each subject. However, in most of the subjects, the process of near to far accommodation was faster than accommodation (P < 0.005, if averaged over all subjects). This correlation was independent of age. 7. The accommodation‐induced pupillary constriction (pupillary near response) was absent in children for a 4 D target; even at 10 D, there was no reliable pupillary response. The pupillary near response increased to about 1.6 mm D‐1 of accommodation at the age of 47. Since a pupillary near response could still be elicited in presbyopic subjects unable to accommodate, the ratio of pupillary constriction per dioptre of accommodation approached infinity. 8. The magnitude of the pupillary near response was highly variable even among subjects of the same age but was typical for each subject. There was a correlation (P < 0.01) to refractive error: corrected myopes had weaker pupillary near responses than emmetropes or hyperopes.(ABSTRACT TRUNCATED AT 400 WORDS)
Afferent and efferent components of the cardiovascular reflex responses to acute hypoxia in term fetal sheep.
Authors: Giussani D A , Spencer J A , Moore P J , Bennet L , Hanson M A
Published: December 2014

1. We studied the effects of acute isocapnic hypoxia on arterial and central venous pressures, carotid and femoral blood flows and heart rate in intact and carotid denervated fetal sheep between 118 and 125 days gestation, after pre‐treatment with either saline, atropine or phentolamine. Electrocortical activity (ECoG) and the incidence of fetal breathing movements (FBM) were also compared between intact and carotid denervated fetuses. 2. There were no significant differences between intact and denervated fetuses in any variable measured during normoxia. Soon after the onset of hypoxia a marked bradycardia occurred in intact, but not in denervated fetuses. Femoral blood flow and femoral vascular resistance (perfusion pressure/femoral blood flow) increased in intact, but not in denervated fetuses. Carotid blood flow increased in both groups of fetuses during hypoxia, but carotid vascular resistance did not change. During hypoxia, the incidence of FBM and low‐voltage ECoG was similarly reduced in both groups of fetuses. 3. Atropine produced a rise in fetal heart rate during the control period in intact but not in denervated fetuses. At the onset of hypoxia atropine prevented the initial bradycardia seen in intact fetuses. In denervated fetuses a further increase in heart rate occurred throughout the hypoxia. 4. All denervated fetuses treated with phentolamine died during the hypoxic challenge, but nine out of fourteen intact fetuses treated with phentolamine survived. 5. In intact fetuses which survived hypoxia after treatment with phentolamine, the increase in arterial blood pressure was smaller and the increase in femoral resistance did not occur. In these fetuses a rise in heart rate occurred in hypoxia. Carotid vascular resistance decreased during hypoxia after administration of phentolamine. 6. Our results indicate that the initial cardiovascular responses of the late gestation sheep fetus to hypoxia are reflex, and that the carotid chemoreceptors provide the afferent limb of this reflex. The bradycardia is mediated through a muscarinic pathway, as it is blocked by atropine. The femoral vasoconstriction is mediated through an alpha‐adrenergic mechanism, mediated both neurally by a carotid chemoreflex and via catecholamines released directly from the adrenal medulla. Both these components are blocked by phentolamine. 7. The differences in survival between intact and denervated fetuses during hypoxia after phentolamine suggest that the carotid chemoreflex response to hypoxia involves mechanisms in addition to vagal efferents to the heart and alpha‐adrenergic actions at peripheral blood vessels.(ABSTRACT TRUNCATED AT 400 WORDS)
Central activation of autonomic effectors during mental simulation of motor actions in man.
Authors: Decety J , Jeannerod M , Durozard D , Baverel G
Published: December 2014

1. Healthy subjects actually performed and mentally simulated a leg exercise at two levels of work (15 and 19 kg loads). Heart rate, respiration rate and end‐tidal PCO2 were measured in both conditions. In addition, muscular metabolism was simultaneously measured using 31P nuclear magnetic resonance (NMR) spectroscopy. 2. During actual exercise, heart and respiration rates increased, first abruptly and then gradually in relation to the level of work. End‐tidal PCO2 was unaltered. NMR spectra showed a drop in phosphocreatine (PCr) and an increase in inorganic phosphate (Pi) concentrations. Intracellular pH fell to 6.65 at maximal effort with a 19 kg load. 3. During mental simulation, both heart and ventilatory rate increased immediately after mental exercise was begun. This increase was proportional to the amount of simulated exercise. Heart rate remained about 25% below the level observed during actual exercise. The increase in respiration rate, by contrast, was more marked than during actual exercise. Finally, end‐tidal PCO2 decreased progressively to about 18% of the resting value. 4. During mental simulation, NMR spectra were unchanged with respect to the resting values. 5. Subjects rated their sensation of fatigue using an analog rating scale, during both actual exercise and mental simulation. During mental exercise, the sensation of fatigue was greater with the 19 kg load than with the 15 kg load. 6. These results demonstrate that mental simulation of action can activate heart and respiration control mechanisms. They suggest that autonomic activation during imagined action pertains to the more general phenomenon of preparation for action.
Functional identities of thoracic respiratory interneurones in the cat.
Authors: Kirkwood P A , Schmid K , Sears T A
Published: December 2014

1. Spike‐triggered averaging was used to reveal focal synaptic potentials (FSPs) in the thoracic ventral horn resulting from impulses in individual respiratory interneurones situated in the contralateral ventral horn of the same segment (T6 or T7) in anaesthetized paralysed cats. 2. FSPs were of two types, negative‐going or positive‐going, and were of the same sign at each of several sites for each unit. These two types were interpreted as indicating excitatory or inhibitory actions respectively. 3. The positive‐going FSPs were almost all the result of activity in the most strongly modulated, phasic respiratory interneurones, whereas most of the negative‐going ones were derived from neurones with a continuous tonic component in their firing patterns. 4. Positive‐going FSPs (particularly those from inspiratory neurones) were generally found at more ventral sites in the ventral horn than negative‐going ones. 5. It is suggested that an important role for the phasically active interneurones is to provide phasic inhibition, which is combined with largely tonic excitation to produce strong respiratory modulation in the activity of other neurones in the thoracic spinal cord, particularly in motoneurones. However, some groups of respiratory interneurones, such as a group of tonic inspiratory ones which are located more dorsally than the phasic inspiratory ones, may not receive such phasic inhibition.
Contribution of the ciliary cyclic nucleotide‐gated conductance to olfactory transduction in the salamander.
Authors: Lowe G , Gold G H
Published: December 2014

1. Flash photolysis of caged cyclic nucleotides was used to examine the contribution of the ciliary cyclic nucleotide‐gated conductance to olfactory transduction in the tiger salamander. Brief illumination of solitary olfactory receptor cells loaded with 100 microM caged cyclic AMP caused a large inward current (peak amplitude 355 +/‐ 200 pA; mean +/‐ S.D. for eleven cells) under whole‐cell voltage clamp at ‐50 mV. 2. The photolysis response was initiated after a latency of 4‐12 ms, whereas an odorant response of identical amplitude had a latency of several hundred milliseconds. The amplitudes of both responses exhibited almost identical voltage dependence between ‐50 and +25 mV, with both reversing near 0 mV. The time courses of the falling phases of odorant and photolysis responses also exhibited similar voltage dependence, both being prolonged at positive voltages. 3. Photolysis of caged cyclic GMP activated a current similar in amplitude and time course to that produced by photolysis of caged cyclic AMP. 4. When the flash was spatially limited to the cilia, the amplitude and duration of the photolysis response increased linearly with the length of the cilia illuminated (for cilia not longer than 30‐40 microns) while the latency remained constant at 4‐12 ms. The increase in duration was described semi‐quantitatively by a model which incorporated diffusion and saturable hydrolysis of cyclic AMP. When the flash was limited to the soma or proximal dendrite, the response latency was proportional to the square of the distance between the illuminated region and the cilia. 5. Dialysis of cells with 500 microM cyclic AMP from a whole‐cell electrode under voltage clamp activated a large transient inward current. Simultaneous suction electrode recording showed that this current originated almost entirely from the ciliary membrane. The density of cyclic nucleotide‐gated channels was estimated to be 800‐fold higher in the cilia than in the soma. 6. Summation of simultaneous odorant and photolysis responses was non‐linear, the flash‐induced current being enhanced during a small odorant response and attenuated during a large odorant response. Summation of two photolysis responses was similarly non‐linear. The data were consistent with odorant stimuli and cyclic AMP both activating a common cyclic nucleotide‐gated conductance with a Hill coefficient, n, of 2.0‐4.4. For n = 2.5, the basal cyclic AMP concentration was estimated to be less than 20% of the K 1/2, which predicts a basal current of 5.8 pA, less than 2% of the maximum.(ABSTRACT TRUNCATED AT 400 WORDS)
The effect of polyamines on voltage‐activated calcium channels in mouse neuroblastoma cells.
Authors: Herman M D , Reuveny E , Narahashi T
Published: December 2014

1. Putrescine has been implicated in modulating cytoplasmic calcium concentration and is correlated with selective neuronal vulnerability in cerebral ischaemia. In order to determine whether putrescine modulates voltage‐activated calcium channels, whole‐cell and single channel patch clamp experiments were performed with N1E‐115 mouse neuroblastoma cells. 2. L‐type calcium channel currents showed a 34 +/‐ 21% increase (n = 6 cells) during external application of 1 mM putrescine. There was no change in the kinetics of the current and no shift in the current‐voltage relationship along the voltage axis. 3. T‐type calcium channel currents were not affected by 1 mM putrescine. 4. The effect of putrescine on single L‐type calcium channels was studied using the cell‐attached configuration of the patch clamp technique. Putrescine (5 mM) applied to the bathing solution, but not present in the pipette, caused an increase in open time of the single channel current without changing the conductance of the channel. In 345 depolarizing steps compiled from three cells, the number of channel openings longer than 3 ms increased from six to seventy‐six, and the number of channel openings longer than 9 ms increased from zero to twenty‐seven. This single channel study supports the hypothesis that putrescine acts on the L‐type channel from the inside of the cell. 5. External application of 1 mM spermine and 1 mM spermidine had no effect on T‐ and L‐type calcium channels. Thus, the effect of putrescine is probably not mediated by the higher polyamines. 6. In order to test whether the effect of putrescine is mediated by a second messenger, specific protein kinase C and cyclic AMP‐dependent protein kinase inhibitors, staurosporine and KT5720, respectively, were applied prior to putrescine. When cells were preconditioned with 200 nM staurosporine, the increase of the L‐type calcium current by 1 mM putrescine was inhibited. By contrast, 200 nM KT5720 did not inhibit the putrescine effect. Therefore, the increase of L‐type channel currents by putrescine may be mediated by protein kinase C but not the cyclic AMP‐dependent protein kinase. 7. The putrescine‐induced enhancement of the L‐type calcium channel activity may play an important role in calcium‐induced neurotoxicity.
Evidence for more than one type of non‐NMDA receptor in outside‐out patches from cerebellar granule cells of the rat.
Authors: Wyllie D J , Traynelis S F , Cull‐Candy S G
Published: December 2014

1. Application of non‐NMDA (non‐N‐methyl‐D‐aspartate) receptor agonists onto outside‐out patches of cerebellar granule cells gave two characteristic types of response (in different patches) which we have referred to as ‘high conductance’ and ‘low conductance’ responses. At a qualitative level these patches could be readily distinguished by the size of the noise increase accompanying their membrane currents. 2. In high conductance patches both AMPA (alpha‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole propionic acid) and kainate gave discrete single‐channel conductances (10‐30 pS), while in low conductance patches, AMPA produced small discrete events (6‐10 pS), and kainate opened channels with conductances too small to be directly resolved. All patches examined contained NMDA receptor channels with characteristic 50 and 40 pS conductance levels. 3. Despite the marked differences in single‐channel conductances, kainate dose‐response curves constructed for high and low conductance patches had similar EC50 values of approximately 150 microM. 4. Spectral analysis of low conductance kainate responses gave an estimated channel conductance of approximately 1.5 pS. In these same low conductance patches AMPA produced discrete openings with two conductance levels; their mean conductances (and relative proportions) were 6 (87%) and 10 pS (13%). 5. In high conductance patches, glutamate (10‐30 microM), AMPA (3‐10 microM), and kainate (10‐30 microM), each activated non‐NMDA channels with three multiple conductance levels. The amplitudes of these conductance levels (approximately 10, 20 and 30 pS) were similar for each of the agonists, and their relative proportions (i.e. areas in the amplitude histograms) were constant for all three agonists. In addition, the relative proportion of levels was constant between patches, and all three levels were invariably present. These observations are all consistent with the idea that the three multiple conductances originate from a single receptor channel, activated by AMPA, kainate and glutamate. 6. Non‐NMDA single‐channel current‐voltage (I‐V) plots showed outward rectification in high conductance patches. For all three multiple conductance levels the ratio of outward to inward single‐channel slope conductance was 1.8 +/‐ 0.1 and this rectification remained present in symmetrical Na+ solutions. 7. In high conductance patches, the events produced by a rapid application of 20‐50 microM glutamate were compared with those activated during steady‐state application.(ABSTRACT TRUNCATED AT 400 WORDS)
Adaptation of cat motoneurons to sustained and intermittent extracellular activation.
Authors: Spielmann J M , Laouris Y , Nordstrom M A , Robinson G A , Reinking R M , Stuart D G
Published: December 2014

1. The main purpose of this study was to quantify the adaptation of spinal motoneurons to sustained and intermittent activation, using an extracellular route of stimulating current application to single test cells, in contrast to an intracellular route, as has been used previously. In addition, associations were tested between firing rate properties of the tested cells and other type (size)‐related properties of these cells and their motor units. 2. Motoneurons supplying the medial gastrocnemius muscle of the deeply anaesthetized cat were stimulated for 240 s with microelectrodes which passed sustained extracellular current at 1.25 times the threshold for repetitive firing. Many cells were also tested following a rest period with intermittent 1 s current pulses (duration 600 ms) at the same relative stimulus strength. Cell discharge was assessed from the EMG of the motor unit innervated by the test neuron. The motoneurons and their motor units were assigned to four categories (i.e. types FF, FR, S and F; where F = FF + FR) based on conventional criteria. In all, twenty F (16 FF, 4 FR) and fourteen S cells were studied with sustained stimulation. Thirty of these cells (17 F, 13 S) and an additional two cells (1 F, 1 S) were studied with intermittent stimulation. 3. The mean threshold current required for sustained firing for a period of > or = 2 s was not significantly different for F and S cells. However, most of the other measured parameters of motoneuron firing differed significantly for these two cell groups. For example, at 1.25 times the threshold current for repetitive firing, the mean firing duration in response to 240 s of sustained activation was 123 +/‐ 88 s (+/‐ S.D.) for F cells vs. 233 +/‐ 19 s for S cells. These values were significantly longer than those from a comparable, previously reported study that employed intracellular stimulation. With intermittent stimulation, the firing durations of F and S cells were not significantly different from each other. 4. All cells exhibited a delay from the onset of current to the first spike, followed by a brief accelerating discharge that was followed by a slower drop in firing rate. Some cells (21 of 34 with sustained activation; 20 of 32 with intermittent) exhibited doublet discharges (interspike intervals < or = 10 ms) that were intermingled with the more predominant singlet discharges. Doublets were more common in the S cell type.(ABSTRACT TRUNCATED AT 400 WORDS)
Muscle stiffness, strength loss, swelling and soreness following exercise‐induced injury in humans.
Authors: Howell J N , Chleboun G , Conatser R
Published: December 2014

1. In order to study injury‐related changes in muscle stiffness, injury to the elbow flexors of thirteen human subjects was induced by a regimen of eccentric exercise. 2. Passive stiffness over an intermediate range of elbow angles was measured with a device which held the relaxed arm of the subject in the horizontal plane and stepped it through the range of elbow angles from 90 deg to near full extension at 180 deg. The relation between static torque and elbow angle was quite linear over the first 50 deg and was taken as stiffness. 3. Stiffness over this range of angles more than doubled immediately after exercise and remained elevated for about 4 days, and may result from low level myofibrillar activation induced by muscle stretch. 4. Arm swelling was biphasic; arm circumference increased by about 3% immediately after exercise, fell back toward normal, then increased by as much as 9% and remained elevated for as long as 9 days. 5. Ultrasound imaging showed most of the swelling immediately following the exercise to be localized to the flexor muscle compartment; subsequent swelling involved other tissue compartments as well. 6. Muscle strength declined by almost 40% after the exercise and recovery was only slight 10 days later; the half‐time of recovery appeared to be as long as 5‐6 weeks.
Effects of nerve growth factor on neuronal plasticity of the kitten visual cortex.
Authors: Carmignoto G , Canella R , Candeo P , Comelli M C , Maffei L
Published: December 2014

1. The effect of intraventricular administration of nerve growth factor (NGF) by means of a cannula‐minipump system was studied in kittens monocularly deprived during the critical period. The ocular dominance of area 17 neurones of NGF‐treated and control kittens was determined by conventional extracellular recordings. The soma size of cells in A and A1 laminae of the lateral geniculate nucleus (LGN) was also evaluated in Cresyl Violet preparations. 2. Binocularly responsive neurons were found to be significantly more numerous in NGF‐treated than in control kittens. The shrinkage of cells from the deprived LGN laminae normally observed in control kittens was prevented by NGF administration. 3. Following an initial period of monocular deprivation (MD) kittens subsequently treated with NGF showed a substantial recovery of functional binocular connections. 4. These findings indicate that the administration of NGF during the period of deprivation reduces the amblyopic effects of MD, while its administration to kittens with both eyes open following the initial deprivation promotes recovery of the deprived eye. 5. Neurotrophic factors may contribute to the regulation of experience‐dependent modifications of synaptic connectivity in the visual cortex.
Unidirectional interaction between two intracellular calcium stores in rat phaeochromocytoma (PC12) cells.
Authors: Reber B F , Stucki J W , Reuter H
Published: December 2014

1. A clone of the rat phaeochromocytoma cell line (PC12) was treated with nerve growth factor (NGF) for 4‐6 days and used to study caffeine‐ and bradykinin‐induced Ca2+ release from intracellular Ca2+ stores. The caffeine‐sensitive store can be depleted by Ca(2+)‐induced Ca2+ release (CICR), while the bradykinin‐induced release is mediated by inositol 1,4,5‐trisphosphate (IP3). The effect of Ca2+ release from these Ca2+ stores on cytosolic free Ca2+ ([Ca2+]i) was measured by means of fura‐2 single cell microfluorimetry. 2. Caffeine application caused no or only a small Ca2+ release in untreated cells in normal culture medium. The caffeine‐sensitive pool could be filled by Ca2+ entry into cells through either voltage‐activated Ca2+ channels or ligand‐gated cation channels. 3. Bradykinin application produced substantial Ca2+ release in untreated cells in normal culture medium. The response was enhanced after K(+)‐depolarization of the cells. The bradykinin‐induced release of Ca2+ also caused depletion of the caffeine‐sensitive pool by CICR. However, Ca2+ released from the IP3‐sensitive store was not sequestered into the caffeine‐sensitive Ca2+ store. 4. The caffeine‐induced rise in [Ca2+]i was blocked by ryanodine in a use‐dependent manner. In addition, a substantial use‐dependent ryanodine block resulted from the bradykinin‐induced rise of [Ca2+]i and subsequent CICR. By contrast, the K(+)‐induced rise of [Ca2+]i caused only a marginal use‐dependent ryanodine inhibition of Ca2+ release. 5. Our results suggest an enhancement of the IP3‐induced [Ca2+]i rise in the cytoplasm by CICR from the caffeine‐sensitive pool. 6. A mathematical model adequately simulates our experimental data.
An in vitro study of the properties of single vagal afferents innervating guinea‐pig airways.
Authors: Fox A J , Barnes P J , Urban L , Dray A
Published: December 2014

1. A novel preparation of the trachea and main bronchi with attached vagus nerve from the guinea‐pig maintained in vitro was used to study the properties of single vagal afferent nerve fibres with identified receptive fields. 2. Recordings were made from twenty‐eight C fibres with a mean conduction velocity of 0.9 +/‐ 0.1 m s‐1 and twenty‐four A delta fibres with a mean conduction velocity of 8.4 +/‐ 1.3 m s‐1. Receptive fields for C and A delta fibres were of small diameter, distributed throughout the trachea and right bronchus and possessed very low mechanical thresholds of 2.2 +/‐ 0.4 and 1.1 +/‐ 0.3 mN respectively. 3. The chemosensitivity of isolated afferents was studied by applying drugs directly onto identified receptive fields. A delta fibres were insensitive to capsaicin (up to 3 microM), bradykinin (3 microM), histamine (10 microM) and 5‐hydroxytryptamine (5‐HT; 10 microM) applied for up to 1 min. Histamine (10 microM), 5‐HT (10 microM) and m‐chlorophenylbiguanide (10 microM) were also ineffective in exciting C fibres. 4. Capsaicin, at concentrations ranging from 30 nM to 3 microM, evoked a sustained firing of all C fibres tested when applied for a period of 30 s directly onto receptive fields. Bradykinin (0.1‐1 microM) also potently excited C fibres in a concentration‐related manner. The effect of bradykinin appeared to be mediated by a B2 receptor since it was not mimicked by the selective B1 receptor agonist [des‐Arg9]‐bradykinin (3 microM) and was abolished by prior application of the selective B2 receptor antagonist D‐Arg‐[Hyp3,Thi5,D‐Tic7,Oic8]‐bradykinin (HOE 140; 0.1 microM). HOE 140 was without effect against capsaicin‐evoked discharge of C fibres. 5. Capsaicin‐ and bradykinin‐evoked discharge of C fibres was present to a similar degree in preparations pretreated with ibuprofen (1 microM), indicating that it was not dependent on, or influenced by, endogenous prostaglandin production. 6. These data demonstrate that single vagal afferents may be studied in vitro and provide the first examination of the properties of sensory fibres innervating guinea‐pig airways. C and A delta fibres both exhibit low threshold mechanical sensitivity but show marked differences in terms of their chemosensitivity.
Brief calcium transients evoked by glutamate receptor agonists in rat dorsal horn neurons: fast kinetics and mechanisms.
Authors: Reichling D B , MacDermott A B
Published: December 2014

1. The calcium indicator dye, indo‐1, was used to analyse the receptor‐specific mechanisms of intracellular calcium ion ([Ca2+]i) responses evoked by excitatory amino acid (EAA) stimulation of dorsal horn neurons. Measurements of somal changes in [Ca2+]i were made on a subsecond time scale under conditions designed to allow membrane potential to mediate interactions between agonist‐gated channels and voltage‐gated calcium channels (VGCCs). 2. Voltage‐gated calcium channels were activated in a receptor‐independent manner using elevated extracellular [K+]. The concentration‐dependence of K(+)‐evoked [Ca2+]i transients was steep and variable among cells, with a mean maximal [Ca2+]i response of 1400 nM and a rapid maximal rate of rise. These data indicate that VGCCs provide a high‐capacity route for Ca2+ entry that is very sensitive to small changes in membrane potential. 3. Stimulation of non‐NMDA receptors using the non‐desensitizing agonist kainate also evoked large [Ca2+]i responses (mean, 840 nM) that were predominantly due to indirect activation of VGCCs. However, in 60% of neurons tested, a component of the [Ca2+]i transient evoked by kainate at concentrations above 10 microM was not blocked by the potent VGCC blocker, lanthanum (La3+). The La(3+)‐resistant [Ca2+]i responses to kainate rose exponentially, required extracellular Ca2+, and were caused neither by evoked release of EAA transmitters nor by reversal of Na(+)‐Ca2+ exchange. These responses may be mediated by a Ca(2+)‐permeable conformation of non‐NMDA receptors and can also be evoked by quisqualate, (S)‐alpha‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole propionic acid (AMPA) and glutamate. 4. Non‐NMDA receptors were activated in a desensitizing manner using quisqualate or AMPA. Quisqualate evoked small [Ca2+]i transients (210 nM) with a slow rate of rise. Typically, above 3 microM quisqualate, the size of the responses decreased, reflecting desensitization of the receptor. Responses to quisqualate were blocked by removal of extracellular Ca2+ indicating that mobilization of intracellular Ca2+ stores does not occur in the majority of dorsal horn neurons. However, trans‐(+‐)‐1‐amino‐1,3‐cyclopentane dicarboxylic acid (trans‐ACPD) was occasionally able to evoke modest Ca2+ release. 5. Activation of the Ca(2+)‐permeable NMDA receptors evoked [Ca2+]i transients that were large (780 nM), with a moderate rate of rise, and that generally achieved a maximum amplitude at NMDA concentrations around 300 microM. 6. Glutamate was used to examine [Ca2+]i responses to the activation of mixed EAA receptor subtypes by an endogenous ligand.(ABSTRACT TRUNCATED AT 400 WORDS)
GABAA and glutamate receptor involvement in dendrodendritic synaptic interactions from salamander olfactory bulb.
Authors: Wellis D P , Kauer J S
Published: December 2014

1. Whole‐cell patch clamp and optical recording techniques were applied to the same in vitro salamander olfactory bulb preparations to study the postsynaptic responses of single mitral/tufted cells in the context of the surrounding neural activity in which they are embedded. Mitral/tufted cells were identified by intracellular filling with biocytin. 2. Single mitral/tufted cells were under a tonic GABAA receptor‐mediated inhibitory influence as revealed by the recording of bicuculline methiodide (BMI)/picrotoxin‐sensitive inhibitory postsynaptic currents (IPSCs) in symmetrical chloride conditions at a holding potential of ‐70 mV. Depolarizing voltage steps (100 ms) applied to single cells or electrical stimulation of the olfactory nerve or medial olfactory tract evoked a prolonged increase in the frequency of GABAergic IPSCs. 3. The frequency of spontaneous and driven IPSCs was reduced with application of the glutamate receptor antagonists 6‐cyano‐2,3‐dihydroxy‐7‐nitro‐quionoxaline (CNQX) or 2‐amino‐5‐phosphonopentanoic acid (AP5) whereas olfactory nerve‐ or medial olfactory tract‐driven IPSC frequency was enhanced with removal of bathing Mg2+, indicating that GABAergic interneurones were driven by mitral/tufted cells at both non‐NMDA and NMDA receptors. 4. Olfactory nerve or medial olfactory tract stimulation evoked widely distributed changes in fluorescence in preparations stained with the voltage‐sensitive dye RH414. The optical response predominantly consisted of a decrease in fluorescence, indicative of depolarization. The presence of the dye did not obviously affect mitral/tufted cell postsynaptic responses. 5. BMI enhanced the amplitude and duration of optical signals related to depolarization within the bulb and in regions central to the bulb. In the presence of BMI, depolarizing activity appeared to spread hundreds of micrometres into regions of the bulb not activated in control conditions showing explicitly that GABAA receptors in the bulb participate in lateral inhibition. 6. CNQX and AP5 attenuated the optical signals within the bulb supporting the contention that in these conditions, optical signals arise mainly from granule cell dendritic activity. Furthermore, AP5 or removal of bath Mg2+ reduced or enlarged the spatial distribution of activity respectively, suggesting that in some cases the NMDA receptor may be involved in generating or stabilizing spatial patterns of activity. 7. It is concluded that in the salamander olfactory bulb, both GABAA‐ and glutamate receptor‐mediated synaptic transmission shape the different temporal and spatial patterns of neural activity associated with olfactory coding.
Characterization of the outward rectifying potassium channel in a novel mouse intestinal smooth muscle cell preparation.
Authors: Molleman A , Thuneberg L , Huizinga J D
Published: December 2014

1. The outward rectifying K+ conductance and underlying single channel behaviour in mouse small intestine (MSI) smooth muscle cells was studied using microelectrode impalement and the patch clamp technique. 2. At 37 degrees C, smooth muscle cells in MSI explants had a resting membrane potential around ‐65 mV and showed spontaneous electrical and mechanical activity. 3. Under whole‐cell voltage clamp, depolarization of smooth muscle cells in the explants evoked a methoxyverapamil (D600)‐sensitive, partially inactivating inward current and a non‐inactivating outward current. The outward current was also observed in enzymatically dispersed cells from neonatal mouse small intestine. 4. The reversal potential of the outward current as established in tail current experiments was ‐70.2 mV. Tail currents could be fitted with a single exponential, suggesting the participation of only one population of channels. 5. The outward current was sensitive to 4‐aminopyridine (10(‐4) M), Ba2+ (1 mM) and to the presence of Cs+ in the pipette, but not to D600 (10(‐6) M), or the presence of ATP (1 mM) in the pipette. 6. In the cell‐attached patch configuration, a unitary outward current was observed that showed increased activity upon depolarization of the patch. The current‐voltage relationship was close to linear with a slope conductance of 186 pS. 7. With normal K+ (6 mM) in the pipette, the extrapolated reversal potential for the unitary current was around ‐75 mV, while with high K+ (120 mM) the reversal potential was close to 0 mV. 8. Averaging single channel traces recorded under a depolarizing pulse protocol resulted in a trace with similar time characteristics as the outward current observed in the whole‐cell configuration. 9. The burst behaviour of the channel was described by a simple model consisting of two closed states, Cf (intraburst closed state) and Cs (interburst closed state) and an open state (O). The rate constants in the model showed differential sensitivity to potential changes, channel blockade by Ba2+ and equimolar K+ conditions. 10. It was concluded that the outward rectifying potassium current in MSI smooth muscle cells is mediated by a 186 pS bursting channel. Voltage dependency and Ba2+ blockade are mainly reflected by changes in the transition rate from the open channel state to the interburst closed state.
Pacemaker potentials in lymphatic smooth muscle of the guinea‐pig mesentery.
Authors: Van Helden D F
Published: December 2014

1. Intracellular microelectrode recordings have been made from lymphatic smooth muscle (SM) to investigate pacemaker mechanisms underlying lymphatic pumping. 2. The SM of small lymphangions or that of short segments, cut from lymphangions of any length, behaved similarly. Both preparations exhibited spontaneous transient depolarizations (STDs) and exhibited simplified electrical characteristics approximating those of a spherical cell. 3. STDs were found to underlie activation of action potentials and hence constrictions. 4. The level of STD activity correlated to the pumping activity of lymphangions, the SM from more active chambers exhibiting increased STD activity. 5. Lymphatic SM exhibited STDs with properties similar to the STDs of mesenteric veins. STDs appeared to be of myogenic origin as they were present despite denervation or substantial destruction of the endothelium. 6. Noradrenaline enhanced the size and frequency of STDs. 7. STD activity was abolished by chelation of cytosolic Ca2+. 8. It is proposed that STDs provide a mechanism for pacemaking in the lymphatic SM studied here. Furthermore, it is postulated that STDs are the consequence of Ca(2+)‐dependent pulsatile release of an intracellular messenger, probably Ca2+ itself. This mechanism provides a novel means for pacemaking.
Energetics of fast‐ and slow‐twitch muscles of the mouse.
Authors: Barclay C J , Constable J K , Gibbs C L
Published: December 2014

1. The energetic cost of work performance by mouse fast‐ and slow‐twitch muscle was assessed by measuring the rates of thermal and mechanical energy liberation of the muscles at 21 degrees C. Thermal energy (heat) liberation was measured using a fast‐responding thermopile. 2. Bundles of muscles fibres from the slow‐twitch soleus and fast‐twitch extensor digitorum longus (EDL) muscles were used. Work output was controlled by performing isovelocity shortenings during the plateau of an isometric tetanus. A range of shortening velocities, spanning the possible range, was used for each muscle. 3. During tetanic contractions, the rate of heat production from EDL muscle was 134.2 +/‐ 11.4 mW/g. The rate of heat production by soleus muscle was only one‐fifth as great (26.8 +/‐ 2.7 mW/g). 4. The maximum shortening velocity (Vmax) of EDL muscles was 2.5‐fold greater than that for soleus muscles and it's force‐velocity relationship was less curved. Peak power output from EDL muscles was 3‐fold greater than that from soleus muscle. 5. During shortening, the rate of heat output from soleus muscles increased considerably above the isometric heat rate. In contrast to soleus muscle, the rate of heat production by EDL muscle increased by only a small fraction of the isometric heat rate. The magnitude of the increases in rate was proportional to shortening velocity. 6. The total rate of energy liberation (heat rate + power) by EDL muscle, shortening at 0.95 Vmax was 1.62 +/‐ 0.37 times greater than the isometric heat rate. In contrast, the rate of energy liberation from soleus muscle shortening at 0.95 Vmax was 5.21 +/‐ 0.58 times greater than its isometric heat rate. The peak mechanical efficiency (power/total energy rate) of the both muscles was approximately 30%.
Dual modulation of K+ currents and cytosolic Ca2+ by the peptide TRH and its derivatives in guinea‐pig septal neurones.
Authors: Toledo‐Aral J , Castellano A , Ureña J , López‐Barneo J
Published: December 2014

1. We describe a dual effect of the peptide TRH (thyrotrophin‐releasing hormone) and its derivatives at concentrations between 0.1 and 1 microM on the K+ currents and cytosolic Ca2+ concentration in enzymatically dispersed septal neurones. 2. In response to membrane depolarization, septal neurones recorded under whole‐cell patch clamp can generate two major K+ currents: (i) a fast and transient K+ current (I(t)), that after a maximum at 2‐5 ms inactivates completely at all membrane potentials in less than 50 ms; and (ii) a slowly activating current (I(s)), which reaches a maximum in 15‐20 ms and does not exhibit appreciable inactivation during short‐lasting voltage pulses. 3. In about 70% of the neurones tested (n = 48) TRH induced a reversible, and often transient, increase of I(t), I(s) or both K+ conductaNces. In approximately 10% of the cells the peptide had an opposite effect and caused a more protracted and partially reversible attenuation of the amplitude of I(t) and I(s). 4. The dual action of TRH on the K+ currents was mimicked by its derivatives but the effects varied depending on their structural relationship with the precursor neuropeptide. The physiological metabolite cyclo‐His‐Pro and the synthetic analogue methyl‐TRH, in which the carboxyl terminus of the molecule is conserved, increased the K+ currents, whereas depression of the K+ conductances was predominantly observed in the presence of TRH‐OH, in which the amino end of TRH is maintained intact. 5. In fura‐2‐loaded unclamped cells, TRH induced either release of Ca2+ from internal stores, Ca2+ entry, or both. With TRH‐OH we never observed mobilization of internal Ca2+ but this peptide evoked a large Ca2+ influx. 6. The results demonstrate that the physiological metabolites of brain TRH (cyclo‐His‐Pro and TRH‐OH) have biological activity. TRH and its derivatives exert two types of regulatory actions on the voltage‐gated K+ channels and cytosolic Ca2+ concentration in central neurones, which can be explained assuming that TRH and TRH‐derived products interact with different subtypes of brain receptors recognizing preferentially either the amino or the carboxyl termini of the TRH molecule.
The role of Na(+)‐Ca2+ exchange in paired pulse potentiation of ferret ventricular muscle.
Authors: Kirby M S , McCall E , Orchard C H , Boyett M R
Published: December 2014

1. Stimulation of cardiac muscle with pairs of stimuli ('paired pulse stimulation') results in a large inotropic effect and experiments have been carried out on ferret ventricular muscle to investigate the underlying mechanism. 2. Aequorin was used to measure sarcoplasmic Ca2+ in papillary muscles. During paired pulse stimulation the first aequorin light transient (i.e. Ca2+ transient) and contraction of the pair increased in amplitude, whereas the second aequorin light transient and contraction were small. When the interval between the pair was decreased, the second aequorin light transient and contraction of the pair were smaller, but the increase in the first aequorin light transient and contraction was greater. 3. The relationship between contraction and the aequorin light transient was the same during paired pulse stimulation and on raising the bathing Ca2+ concentration. It is concluded that there was no change in the myofilament sensitivity to Ca2+ during paired pulse stimulation. 4. The increase in the aequorin light transient and contraction during paired pulse stimulation was prevented by ryanodine, an inhibitor of the sarcoplasmic reticulum (SR). 5. During paired pulse stimulation of ventricular myocytes there was little change in the first action potential of the pair, but the second action potential was shorter than control when the interval between the pair was short. During paired pulse stimulation of ventricular myocytes under voltage clamp control there was little change in the first Ca2+ current (iCa) of the pair, but the second iCa was smaller than control when the interval between the pair was short. Because paired pulse potentiation was greatest when the interval between the pair was short, it is concluded that paired pulse potentiation was not the result of a prolongation of the action potential or increase in iCa. 6. During paired pulse stimulation of ventricular myocytes under voltage clamp control the increase in contraction was greater, the more positive the membrane potential during the second pulse of the pair. This voltage dependence is consistent with a role for the Na(+)‐Ca2+ exchanger in paired pulse potentiation. 7. During paired pulse stimulation of ventricular myocytes under voltage clamp control, changes in putative Na(+)‐Ca2+ exchange current were observed consistent with a decrease of Ca2+ efflux (or increase of Ca2+ influx) via the exchanger during the second pulse of the pair. 8. A computer model of excitation‐contraction coupling (Harrison, McCall & Boyett, 1992) has been used to simulate paired pulse stimulation and the results described above.(ABSTRACT TRUNCATED AT 400 WORDS)
Segmental effects of epidural spinal cord stimulation in humans.
Authors: Hunter J P , Ashby P
Published: December 2014

1. The segmental effects of spinal cord stimulation (SCS) were studied in twenty‐four human subjects who had spinal cord stimulators implanted for the treatment of pain. The cathode was in the epidural space over the dorsum of the thoracic cord. 2. SCS generated action potentials in sensory, motor and mixed nerves which could be recorded with near‐nerve electrodes. These action potentials could follow high frequencies of stimulation and appeared to be due to the antidromic activation of primary afferents in the dorsal columns. 3. Synaptic actions on single lumbosacral motoneurons were derived from peristimulus time histograms (PSTHs) of single motor units. SCS produced a brief short‐latency period of increased firing probability (PIF) in motoneurons of all of the muscles examined, probably representing monosynaptic activation. It is argued that the facilitation arises from the antidromic activation of Ia afferents in the dorsal columns. This is the probable explanation for the muscle contractions that can be induced by SCS. 4. SCS inhibited short‐latency group I homonymous facilitation and reciprocal inhibition. The mechanism appears to be presynaptic to the motoneurons and may represent collision in Ia afferents, presynaptic inhibition or homosynaptic depression. 5. It was difficult to demonstrate consistent effects of SCS on reflex pathways from cutaneous afferents to flexor motoneurons because the effects of stimulation of cutaneous nerves on these motoneurons were themselves variable. 6. It is concluded that SCS applied with epidural electrodes over the dorsal cord activates primary afferents in the dorsal columns. Antidromic activation of these afferents results in strong monosynaptic facilitation of motoneurons as well as reduction in transmission in some reflex pathways to motoneurons.