bluuebunny/crossref_metadata_2025 huggingface dataset:
Influence of atrial natriuretic factor on fluid efflux from the splenic circulation of the rat.
Authors: Deng Y , Kaufman S
Published: December 2014

1. Atrial natriuretic factor (ANF) causes a reduction in plasma volume that is abolished by splenectomy. Experiments were conceived to investigate whether ANF acts within the spleen to increase efflux of fluid from the intravascular to the extravascular space. 2. ANF, infused into the splenic artery of anaesthetized rats at rates of 1, 5 and 20 ng min ‐1, caused a dose‐dependent increase in the arteriovenous difference in haematocrit as blood flowed through the spleen (basal difference, 0.18 +/‐ 0.10%; difference after 10 min at 20 ng min ‐1 ANF, 1.5 +/‐ 0.18%; n = 6). There was no such change in plasma protein concentration. 3. ANF (20 ng min ‐1) did not alter splenic arterial blood flow. However, splenic venous blood flow fell so that the arteriovenous difference increased significantly (basal difference, 0.34 +/‐ 0.19 ml min ‐1; difference at 60 min, 1.1 +/‐ 0.20 ml min‐1, n = 7). There was no change in mean arterial pressure. 4. These data confirm our hypothesis that ANF acts within the spleen to increase fluid efflux from the intravascular to the extravascular space. Since there is no change in total splenic blood flow, we propose that the effects of ANF are mediated by dilatation of the splenic afferent arterioles and constriction of the efferent venules, thus increasing filtration pressure.
Basal responses of the L‐type Ca2+ and hyperpolarization‐activated currents to autonomic agonists in the rabbit sino‐atrial node.
Authors: Zaza A , Robinson R B , DiFrancesco D
Published: December 2014

1. The dose dependence of the cholinergic agonist acetylcholine (ACh) and the beta‐adrenergic agonist isoprenaline (Iso) were determined for the hyperpolarization‐activated current (If) and the L‐type Ca2+ current (ICa,L) in single cells isolated from the rabbit sino‐atrial (SA) node. 2. ACh inhibited If by a negative shift of its activation curve with a maximal effect of ‐9.9 mV; half‐maximal effect was produced by 0.019 microM ACh. High ACh concentrations were required to inhibit ICa,L only partially (31% inhibition at 300 microM). 3. In contrast, If and ICa,L responded to Iso over a similar dose range, with concentrations for half‐maximal enhancement of 0.0136 and 0.0070 microM, respectively. 4. The effects on spontaneous activity of ACh (range 0.001‐0.03 microM) and Iso (range 0.001‐1 microM) were investigated. ACh decreased the slope of diastolic depolarization at concentrations similar to those inhibiting If (> 50% at 0.03 microM). Iso enhanced diastolic depolarization at concentrations similar to those affecting both If and ICa,L (half‐maximal effect at 0.027 microM). 5. In a ramp‐clamp protocol simulating diastolic depolarization, the threshold for activation of inward nifedipine‐sensitive current was ‐41.22 +/‐ 0.68 mV. Although enhancing ICa,L, Iso did not affect this threshold. 6. Half‐maximal ACh concentrations for inhibition of automaticity and If are similar and are lower than the threshold concentrations for modulation of ICa,L; this argues against a role of ICa,L in direct muscarinic modulation of pacemaking. In contrast, modulation of If, ICa,L and automaticity occur at similar Iso concentrations. The difference between maximum diastolic potential (‐61.95 +/‐ 0.93 mV) and the threshold for Iso‐stimulated ICa,L (‐39.54 +/‐ 1.03 mV) suggests that this current plays a role only at later stages of diastolic depolarization.
Effects of immunization against an autocrine inhibitor of milk secretion in lactating goats.
Authors: Wilde C J , Addey C V , Peaker M
Published: December 2014

1. Lactating goats were immunized against the goat's milk protein identified as a feedback inhibitor of lactation (FIL). Immunization was by three treatments during the declining stage of lactation. 2. When antibodies to FIL were consistently detected in milk (in response to the third treatment), the rate of decline in milk secretion was significantly reduced compared with sham‐immunized controls. Such a response was not apparent with the first two treatments when serum but not milk titres of anti‐FIL were raised. 3. When one gland of immunized goats was switched (after the third immunization) from twice‐ to once‐daily milking, the ipsilateral decrease in the rate of milk secretion was reduced significantly compared with sham‐immunized goats. 4. The results are compatible with the hypothesis that the autocrine agent FIL acts during milk accumulation as an inhibitor of milk secretion.
Immunomodulation of afferent neurons in guinea‐pig isolated airway.
Authors: Riccio M M , Myers A C , Undem B J
Published: December 2014

1. The trachea, larynx and main bronchi with the right vagus nerve and nodose ganglion were isolated from guinea‐pigs passively immunized 24 h previously with serum containing anti‐ovalbumin antibody. 2. The airways were placed in one compartment of a Perspex chamber for recording of isometric tension while the nodose ganglion and attached vagus nerve were pulled into another compartment. Action potentials arriving from single airway afferent nerve endings were monitored extracellularly using a glass microelectrode positioned near neuronal cell bodies in the ganglion. Mechanosensitivity of the nerve endings was quantified using calibrated von Frey filaments immediately before and after exposure to antigen (10 micrograms ml‐1 ovalbumin). 3. Ten endings responded to the force exerted by the lowest filament (0.078 mN) and were not further investigated. In airways from thirteen immunized guinea‐pigs, the mechanical sensitivity of A delta afferent fibres (conduction velocity = 4.3 +/‐ 0.6 m s‐1) was enhanced 4.1 +/‐ 0.9‐fold following airway exposure to antigen (P < 0.005). Mechanical sensitivities of afferent fibres (conduction velocity = 4.3 +/‐ 0.6 m s‐1) from non‐immunized control guinea‐pig airways were unaffected by antigen (n = 13). 4. Antigen did not overtly cause action potential generation except in one instance when the receptive field was located over the smooth muscle. This ending also responded to methacholine suggesting that spatial changes in the receptive field, induced by muscle contraction, were responsible for the activation. 5. The mediators responsible for these effects are unknown, although histamine, prostaglandins, leukotrienes and tachykinins do not appear to be essential. The increase in mechanical responsiveness was not associated with the smooth muscle contraction since leukotriene C4, histamine and tachykinins, which all caused a similar contraction to antigen, did not affect mechanical thresholds. Moreover, the antigen‐induced increases in excitability persisted beyond the duration of the smooth muscle contraction. 6. These results demonstrate that antigen‐antibody‐mediated inflammatory processes may enhance the excitability of vagal afferent nerve terminals projecting from the airway and thus may contribute to the pathophysiology of allergic airway diseases.
Plasma catecholamines and hyperglycaemia influence thermoregulation in man during prolonged exercise in the heat.
Authors: Mora-Rodríguez R , González-Alonso J , Below P R , Coyle E F
Published: December 2014

1. We manipulated plasma catecholamines (combined adrenaline and noradrenaline concentrations) to three levels during prolonged exercise to determine their effect on cutaneous and forearm vascular conductance (CVC and FVC), oesophageal temperature (T(oes)) and cardiovascular responses. 2. On three occasions, seven endurance‐trained men cycled at 65% VO2, max in the heat (33.1 +/‐ 0.7 degrees C) for 120‐150 min. During the control trial (150 min duration), 0.45% saline was intravenously infused (SI) starting at 30 min, at a rate that replaced a third of the fluid losses. The infusion start time and rate were identical in all three trials. During SI, plasma catecholamine levels increased progressively and were 18.2 +/‐ 2.7 pmol ml‐1 at 150 min. In another trial (120 min duration), adrenaline was infused (AI) at 0.1 microgram kg‐1 min‐1 and plasma catecholamine levels were elevated 6 pmol ml‐1 above SI during the 60‐120 min period. In a third trial (150 min duration), an 18% glucose solution was infused (GI) at a rate that maintained plasma glucose levels above 11 mM and plasma catecholamine levels were 5.0‐5.5 pmol ml‐1 lower (P < 0.05) than SI from 120‐150 min. 3. Heat production and sweat rate were not different during the three trials and neither was the decline in stroke volume, cardiac output and mean arterial pressure. 4. Soon after beginning AI, CVC decreased 15%, T(oes) increased by 0.4 +/‐ 0.1 degree C and heart rate increased by 6 +/‐ 1 beats min‐1; these significant (P < 0.05) differences from SI were maintained throughout the bout. As a result of GI, FVC was 15% higher than SI and T(oes) and heart rate were attenuated by 0.3 +/‐ 0.1 degree C and 7 +/‐ 1 beats min‐1 at 150 min compared with SI (P < 0.05). 5. In conclusion, large increases in plasma catecholamine levels cause hyperthermia during exercise by vasoconstricting the skin. The mechanisms by which hyperglycaemia (i.e. 11 mM) attenuates hyperthermia are less clear and may be due to others factors besides attenuation of the plasma catecholamine response to exercise.
Distribution of neurones expressing inwardly rectifying and Ca(2+)‐permeable AMPA receptors in rat hippocampal slices.
Authors: Isa T , Itazawa S , Iino M , Tsuzuki K , Ozawa S
Published: December 2014

1. Current‐voltage (I‐V) relationships and Ca2+ permeability of receptor channels activated by bath application of kainate, a non‐desensitizing agonist of alpha‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA) receptors, were examined in various types of neurones in hippocampal slices of 5‐ to 13‐day‐old rats by using the tight‐seal patch clamp recording technique. 2. Three types of responses were observed: type I response with outwardly rectifying I‐V relationship, type II response with I‐V relationship of marked inward rectification, and intermediate response with I‐V relationship of weaker inward rectification. Neurones with type I, type II and intermediate I‐V relationships of kainate responses were referred to as type I, type II and intermediate neurones, respectively. 3. Permeability of Ca2+ ions was estimated by the reversal potential of kainate response in the outside‐out patch in Na(+)‐free extracellular solution containing 100 mM Ca2+. The reversal potentials were ‐44.4 +/‐ 14.0 mV (mean +/‐ S.D.) for type I (n = 7), +11.8 +/‐ 3.6 mV for type II (n = 5), and ‐8.7 +/‐ 7.4 mV for the intermediate neurones (n = 7). The values of PCa/PCs, the ratios of the permeability coefficients of Ca2+ and Cs+, estimated according to the constant‐field equation were 0.08 for type I, 1.71 for type II, and 0.50 for the intermediate neurones. 4. Type II and intermediate responses were observed mainly in non‐pyramidal neurones in various areas of the hippocampus, most frequently observed in the stratum molecular of the dentate gyrus and in the stratum radiatum and the stratum lacunosum‐molecular of both the CA1 and CA3 regions. Both type II and intermediate neurones stained with biocytin had round‐ or ellipsoidal‐shaped somata and issued divergent axonal projections to the surrounding structures. 5. Excitatory postsynaptic currents (EPSCs) recorded in type II neurones had 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX)‐sensitive fast and D‐2‐amino‐5‐phosphonovalerate (APV)‐sensitive slow components. The I‐V relationship of the fast component showed a strong inward rectification, indicating that inwardly rectifying AMPA receptors are involved in excitatory synaptic transmission.
The role of the A(2A) adenosine receptor subtype in functional hyperaemia in the hindlimb of anaesthetized cats.
Authors: Poucher S M
Published: December 2014

1. The present study was designed to investigate the contribution of the A(2A) adenosine receptor subtype in the functional hyperaemia response during muscle contraction. 2. In cats anaesthetized with sodium pentobarbitone and breathing spontaneously following tracheotomy, the left sciatic and femoral nerves were electrically stimulated at 3 Hz for 20 min to induce muscle contraction, and hindlimb blood flow was measured with a flow probe. The contribution of the A(2A) adenosine receptor subtype was assessed using ZM 241385, a potent and selective A(2A) adenosine receptor antagonist. 3. In a control group, the muscle isometric tension measured in the extensor digitorum longus‐tibialis anterior muscle group was 6.64 +/‐ 0.66 kg (100 g muscle mass)(‐1) and hindlimb vascular conductance was 0.22 +/‐ 0.03 ml mmHg(‐1)(kg body mass)(‐1) at 20 min of contraction. Administration of vehicle did not affect these parameters upon a second contraction period: 6.31 +/‐ 0.61 kg (100 g muscle mass)(‐1) and 0.23 +/‐ 0.03 ml mmHg(‐1) (kg body mass)(‐1), respectively. Total hindlimb conductance during contraction was unaffected (5.5 +/‐ 3.7% decrease). 4. ZM 241385 (1.0 mg kg(‐1)) did not alter the amount of force produced by the muscle at 20 min of contraction. Hindlimb conductance response was reduced by 27.1 +/‐ 4.8% following the A(2A) selective adenosine receptor antagonist, similar to that observed with the non‐selective antagonist 8‐phenyltheophylline. 5. These results show that adenosine acting at the A(2A) subtype receptor can contribute up to 30% of the functional hyperaemia response in the hindlimb of anaesthetized cats.
Regional differences in the negative inotropic effect of acetylcholine within the canine ventricle.
Authors: Yang Z K , Boyett M R , Janvier N C , McMorn S O , Shui Z , Karim F
Published: December 2014

1. Regional differences in the effects of ACh on sub‐epicardial, mid‐wall and sub‐endocardial cells of the dog left ventricle have been studied. 2. ACh produced a dose‐dependent, atropine‐sensitive negative inotropic effect that was greatest in sub‐epicardial cells and small or absent in sub‐endocardial cells. 3. In sub‐epicardial (but not sub‐endocardial) cells, ACh also resulted in a dose‐dependent decrease in action potential duration. The inotropic effect of ACh on sub‐epicardial cells was primarily the result of the decrease of action potential duration, because during trains of voltage clamp pulses the inotropic effect of ACh was reduced or abolished. At a holding potential of ‐80 mV, 10(‐5)M ACh decreased L‐type Ca2+ current by approximately 8% and this is thought to be responsible for the small inotropic effect during trains of pulses. 4. Although 4‐AP, a blocker of the transient outward current (I(to)), abolished the ‘spike and dome’ morphology of the sub‐epicardial action potential, it had little or no effect on the actions of ACh on sub‐epicardial cells. ACh had no effect on I(to) in sub‐epicardial cells in voltage clamp experiments. 5. ACh activated a Ba(2+)‐sensitive outward current (IK,ACh) in sub‐epicardial cells, but little or no such current in sub‐endocardial cells. In sub‐epicardial cells, ACh also inhibited the inward rectifier current, IK,1. 6. It is concluded that in left ventricular sub‐epicardial cells, ACh activates IK,ACh. This results in a shortening of the action potential and, therefore, a negative inotropic effect. In subendocardial cells, ACh activates little or no IK,ACh and, therefore, it has little or no negative inotropic effect. This may result from a regional variation in the expression of the muscarinic K+ channel.
Asynchronous transmitter release: control of exocytosis and endocytosis at the salamander rod synapse.
Authors: Rieke F , Schwartz E A
Published: December 2014

1. We have studied exocytosis and endocytosis in the synaptic terminal of salamander rods using a combination of Ca2+ imaging, capacitance measurement and the photolysis of Ca2+ buffers. 2. The average cytoplasmic Ca2+ concentration at the dark resting potential was 2‐4 microM. 3. An average cytoplasmic Ca2+ concentration of 2‐4 microM maintained a high rate of continuous exocytosis and endocytosis. 4. Changes in the rate of exocytosis were followed in less than 0.7 s by compensatory changes in the rate of endocytosis. 5. Vesicle cycling in the rod synapse is specialized for graded transmission and differs from that previously described for synapses that release synchronized bursts of transmitter.
Training affects the development of postural adjustments in sitting infants.
Authors: Hadders-Algra M , Brogren E , Forssberg H
Published: December 2014

1. The present study addressed the question of whether daily balance training can affect the development of postural adjustments in sitting infants. 2. Postural responses during sitting on a moveable platform were assessed in twenty healthy infants at 5‐6, 7‐8 and 9‐10 months of age. Multiple surface EMGs and kinematics were recorded while the infants were exposed to slow and fast horizontal forward (Fw) and backward (Bw) displacements of the platform. After the first session the parents of nine infants trained their child's sitting balance daily. 3. At the youngest age, when none of the infants could sit independently, the muscle activation patterns were direction specific and showed a large variation. This variation decreased with increasing age, resulting in selection of the most complete responses. Training facilitated response selection both during Fw and Bw translations. This suggests a training effect on the first level of the central pattern generator (CPG) model of postural control. 4. Training also affected the development of response modulation during Fw translations. It accelerated the development of: (1) the ability to modulate EMG amplitude with respect to platform velocity and initial sitting position, (2) antagonist activity and (3) a distal onset of the response. These findings point to a training effect on the second level of the CPG model of postural adjustments.
Functional characterization of Kv channel beta‐subunits from rat brain.
Authors: Heinemann S H , Rettig J , Graack H R , Pongs O
Published: December 2014

1. The potassium channel beta‐subunit from rat brain, Kv beta 1.1, is known to induce inactivation of the delayed rectifier channel Kv1.1 and Kv1.4 delta 1‐110. 2. Kv beta 1.1 was co‐expressed in Xenopus oocytes with various other potassium channel alpha‐subunits. Kv beta 1.1 induced inactivation in members of the Kv1 subfamily with the exception of Kv 1.6; no inactivation of Kv 2.1, Kv 3.4 delta 2‐28 and Kv4.1 channels could be observed. 3. The second member of the beta‐subunit subfamily, Kv beta 2, had a shorter N‐terminal end, accelerated inactivation of the A‐type channel Kv 1.4, but did not induce inactivation when co‐expressed with delayed rectifiers of the Kv1 channel family. 4. To test whether this subunit co‐assembles with Kv alpha‐subunits, the N‐terminal inactivating domains of Kv beta 1.1 and Kv beta 3 were spliced to the N‐terminus of Kv beta 2. The chimaeric beta‐subunits (beta 1/ beta 2 and beta 3/ beta 2) induced fast inactivation of several Kv1 channels, indicating that Kv beta 2 associates with these alpha‐subunits. No inactivation was induced in Kv 1.3, Kv 1.6, Kv2.1 and Kv3.4 delta 2‐28 channels. 5. Kv beta 2 caused a voltage shift in the activation threshold of Kv1.5 of about ‐10 mV, indicating a putative physiological role. Kv beta 2 had a smaller effect on Kv 1.1 channels. 6. Kv beta 2 accelerated the activation time course of Kv1.5 but had no marked effect on channel deactivation.
NO decreases evoked quantal ACh release at a synapse of Aplysia by a mechanism independent of Ca2+ influx and protein kinase G.
Authors: Mothet J P , Fossier P , Tauc L , Baux G
Published: December 2014

1. The exogenous nitric oxide (NO) donor, SIN‐1, decreased the postsynaptic response evoked by a presynaptic spike at an identified cholinergic neuro‐neuronal synapse in the buccal ganglion of Aplysia californica. 2. The statistical analysis of long duration postsynaptic responses evoked by square depolarizations of the voltage‐clamped presynaptic neurone showed that the number of evoked acetylcholine (ACh) quanta released was decreased by SIN‐1, pointing to a presynaptic action of the drug. 3. Vitamin E, a scavenger of free radicals, prevented the effects of SIN‐1 on ACh release. SIN‐1 still decreased ACh release in the presence of superoxide dismutase, whereas haemoglobin suppressed the effects of SIN‐1. These results showed that NO is the active compound. 4. 8‐Bromoguanosine 3', 5' cyclic monophosphate (8‐Br‐cGMP) mimicked the inhibitory effect of NO on ACh release suggesting the involvement of a NO‐sensitive guanylate cyclase. This was reinforced by the reversibility of the effects of SIN‐1 by inhibitors of guanylate cyclase, Methylene Blue, cystamine or LY83583. Methylene Blue partially reduced the inhibitory effect of NO. In addition, in the presence of superoxide dismutase, Methylene Blue blocked and cystamine significantly reduced the NO‐induced inhibition of ACh release. 5. In the presence of KT5823 or R‐p‐8‐pCPT‐cGMPS, two inhibitors of protein kinase G, the reduction of ACh release by SIN‐1 still took place indicating that the effects of NO most probably did not involve protein kinase G‐dependent phosphorylation. 6. Presynaptic voltage‐dependent Ca2+ (L‐, N‐ and P‐types) and K+ (IA and late outward rectifier) currents were unmodified by SIN‐1. 7. The modulation of ACh release in opposite ways by L‐arginine and N omega‐nitro‐L‐arginine points to the involvement of an endogenous NO synthase‐dependent regulation of transmitter release.
Dynamic modulation of excitation‐contraction coupling by protein phosphatases in rat ventricular myocytes.
Authors: duBell W H , Lederer W J , Rogers T B
Published: December 2014

1. The effects of the serine/threonine protein phosphatases (PP) type 1 and 2A on L‐type Ca2+ current (ICa) and the intracellular [Ca2+]i transient were examined in rat ventricular myocytes. ICa was measured under voltage clamp using patch‐type microelectrodes in the whole‐cell mode with the cells in a steady state of sarcoplasmic reticulum (SR) Ca2+ loading. [Ca2+]i transients were measured simultaneously using the fluorescent Ca2+ indicator indo‐1 (50 microM) which was added to the pipette filling solution along with PP‐1 or PP‐2A (4 units ml‐1). 2. PP‐1 had no effect on the ICa‐V relationship but decreased the [Ca2+]i‐voltage relationship (by 43% at 0 mV). PP‐2A decreased both ICa‐V (by 26% at 0 mV) and the [Ca2+]i transient‐voltage (by 65% at 0 mV). Excitation‐contraction coupling gain, defined as (delta [Ca2+]i/ICa), was decreased to 43% of control by PP‐1 and to 29% of control by PP‐2A at‐28 mV. 3. Diastolic [Ca2+]i (i.e.[Ca2+]i measured immediately before each voltage clamp pulse) was not altered by PP‐1 or PP‐2A and neither phosphatase changed steady‐state SR Ca2+ content, as measured with caffeine. 4. We conclude that the reduced [Ca2+]i transient following the application of PP‐1 was due to reduced SR Ca2+ release channel activity. The effects of PP‐2A, while more broadly based, were still consistent with a decrease in SR Ca2+ release channel activity. 5. Our experiments, combined with recent experiments by others, suggest that the basal state of contractility in heart is dynamically regulated by dephosphorylation and phosphorylation of the SR Ca2+ release channel.
Modulation by purines of calcium‐activated non‐selective cation channels in the outer hair cells of the guinea‐pig cochlea.
Authors: Van den Abbeele T , Tran Ba Huy P , Teulon J
Published: December 2014

1. The cell‐attached and cell‐free configurations of the patch‐clamp technique were used to investigate whether external ATP and its derivatives modulate channel activity in outer hair cells freshly isolated from the guinea‐pig cochlea. 2. Submicromolar concentrations of ATP stimulated a non‐selective cation channel with a conductance of about 25 pS. The ATP‐elicited stimulation was partly blocked by the membrane‐permeant blocker 3',5‐dichlorodiphenylamine‐2‐carboxylic acid (DCDPC), and mimicked by the calcium ionophore, ionomycin, suggesting that the channel activated by ATP is identical to a previously reported calcium‐activated non‐selective (CAN) cation channel. 3. The P2x agonist beta, gamma‐methylene‐ATP (beta, gamma‐MeATP, 10 microM) and the P2Y agonist 2‐methyl‐thio‐ATP (2‐MeSATP, 1 microM) both activated CAN channels. The effect of ATP was inhibited by the P2 antagonist suramin but not by the P2Y antagonist Reactive Blue 2. These results suggest that both purinergic receptors are involved in the ATP‐evoked response and that internal calcium acts as a second messenger for opening CAN channels. 4. In contrast, adenosine inhibited CAN channels. This effect was reproduced by the A2 agonist 5'‐N‐ethylcarboxyamidoadenosine (NECA) and the permeant cAMP analogue 8‐bromo‐adenosine 3',5'‐cyclic monophosphate (8‐Br‐cAMP), but not by the A1 agonist N6‐cyclo‐hexyladenosine (CHA). CAN channels were also inhibited when the catalytic subunit of protein kinase A was applied internally on inside‐out patches, suggesting that adenosine A2 receptor downregulates CAN channels via a cAMP‐dependent phosphorylation.
Voltage dependence of mouse acetylcholine receptor gating: different charge movements in di‐, mono‐ and unliganded receptors.
Authors: Auerbach A , Sigurdson W , Chen J , Akk G
Published: December 2014

1. The voltage dependence of binding and gating in wild‐type and mutant recombinant mouse nicotinic acetylcholine receptors (AChRs) was examined at the single‐channel level. 2. The closing rate constant of diliganded receptors decreased e‐fold with approximately 66 mV hyperpolarization in both wild‐type (adult and embryonic) and mutant receptors. The opening rate constant of a mutant receptor (alpha Y93F) was not voltage dependent. 3. The voltage dependence of closing in monoliganded receptors was examined in several receptors having a mutation in the binding site (alpha G153S) or pore region (alpha L251C and epsilon T264P). The closing rate constant of these monoliganded receptors decreased e‐fold with approximately 124 mV hyperpolarization. 4. The voltage dependence of closing and opening in unliganded receptors was examined in two receptors having a mutation in the pore region (alpha L251C and epsilon T264P). Neither the closing nor the opening rate constants of unliganded receptors were voltage dependent. 5. If z if the amount of charge that moves during channel closure and delta is the distance (as a fraction of the electric field) that the charge moves, we conclude that z delta = 0.4 in diliganded receptors, 0.2 in monoliganded receptors, and 0.0 in unliganded receptors. It is likely that charges on the protein, rather than the agonist molecule, move z delta = 0.2 after each ACh molecule has bound. 6. The results suggest that unliganded openings arise from a local, concerted change in the structure of the pore (channel opening) that does not involve the net movement of charged residues. We speculate that as a consequence of agonist binding, charged moieties in the protein change their disposition so that they move with respect to the electric field when the channel gates. The results are consistent with the idea that there is semi‐independent movement of distinct domains during AChR gating.
Parallel organization of proprioceptive inputs from joint receptors to cortical somatosensory areas I and II in the cat.
Authors: Mackie P D , Zhang H Q , Schmidt R F , Rowe M J
Published: December 2014

1. Studies in monkeys indicate that proprioceptive and tactile inputs are conveyed from the thalamus to the primary somatosensory cortex (SI) and thence to the secondary somatosensory area (SII) in a serial scheme. In contrast, in the cat, tactile information is conveyed in parallel from the thalamus to SI and SII. The present study, in the cat, employed reversible inactivation of SI to determine whether proprioceptive inputs to SII from joint receptors depend on an indirect serial path via SI or are conveyed over a direct path from the thalamus. 2. SI and SII foci for knee joint inputs were determined with evoked potential mapping. Reversible inactivation of the SI focus by cooling had no effect on the amplitude, latency or time course of SII potentials evoked by joint inputs. There was also no consistent effect on the response levels of individual SII neurones examined during SI inactivation. Furthermore, there was no attenuation of the later components of the responses, and therefore no evidence that these depended on an indirect path to SII via SI. 3. Results demonstrate that proprioceptive inputs project directly from thalamus to SII over a pathway organized in parallel with that to SI, in contrast to the serial scheme reported for proprioceptive processing in primates.
External‐anion‐dependent anionic current in blastoderm cells of early medaka fish embryos.
Authors: Shigemoto T , Okada Y
Published: December 2014

1. Anionic current was studied by a whole‐cell variation of the patch‐clamp technique in blastoderm cells dissociated from medaka (Oryzias latipes) embryos at the early blastula stage. The blastoderm cells were mechanically dissociated without using proteolytic enzymes. 2. The anionic current was deactivated by hyperpolarizing steps. The steady‐state current‐voltage (I‐V) relationship of the anion current was accurately represented by the Boltzmann relation with z = 1.01 +/‐ 0.02 (+/‐ S.E.M., n = 4) for voltage‐dependent activation when internal calcium ions were buffered at 100 nM by BAPTA‐Ca2+ buffer. 3. When the internal calcium concentration was reduced to 10 nM, this anionic current became an external‐calcium‐dependent current and was remarkably decreased by removal of external calcium ions. Furthermore, this anionic current was almost abolished when the internal calcium concentration was greatly reduced by chelating Ca2+ ions with 10 mM BAPTA. 4. This current was not affected by replacing external Na+ with TEA+ or N‐methyl‐D‐glucamine. The reversal potentials shifted by +58.6 mV for a 10‐fold decrease in the external Cl‐concentration. External Cl‐ions were substituted with various anions and respective current‐reversal potentials were measured. The order of permeability was I‐ > Br‐ > Cl‐ > F‐. Both the outward and inward currents almost disappeared in external Cl(‐)‐free solutions. 5. The voltage dependency of the anionic current shifted in a positive direction with the reduction in the external Cl‐ concentration, the potentials at which half the channels were activated (V 1/2) being ‐59.8 +/‐ 1.2 mV (n = 4), ‐40.7 +/‐ 0.8 mV (n = 4), ‐14.9 +/‐ 0.7 mV (n = 4), and ‐0.7 +/‐ 0.6 mV (n = 4) for 135, 90, 67, and 45 mM Cl‐, respectively. This implies that the inward current, brought by efflux of internal anions, decreases with reductions in the external Cl‐ concentration. In the presence of external 135 mM Br‐, the voltage dependency of the anionic current shifted in the negative direction (V 1/2 = ‐84.5 +/‐ 1.0 mV and z = 0.91 +/‐ 0.02 (n = 3)) in comparison with that in the external Cl‐ medium. It further shifted in the negative direction in the presence of 135 mM I‐ (V 1/2 = ‐93.8 +/‐ 1.5 mV and z = 0.85 +/‐ 0.03 (n = 3)). The inward current of the anion channel is considered to be affected by both the type and the concentration of external halide ions. 6. At the beginning of the blastula stage, the blastocoel cavity is formed for the first time. A mechanism is necessary to transport ions to the cavity without losing them to pond water outside the embryos. Since this anion channel is external‐anion dependent, the current cannot be activated in the membrane facing the pond water, where concentrations of anions are much lower than those of the intracavernous solution. Only after blastocoel formation are blastoderm cells first exposed to the extracellular fluid with high ionic concentrations. Then, the anion channel would become effective for maintaining the Cl‐ equilibrium potential.
Increased neuronal excitability during depolarization‐induced suppression of inhibition in rat hippocampus.
Authors: Wagner J J , Alger B E
Published: December 2014

1. Depolarization‐induced suppression of inhibition (DSI) is a form of plasticity of gamma‐amino‐butyric acid (GABAA)‐mediated (henceforth ‘GABAergic’) responses in the CNS. We made whole‐cell recordings from CA1 pyramidal neurons to investigate the effects of DSI on excitatory synaptic transmission in the hippocampal slice preparation. 2. Significant enhancement of the voltage‐clamped excitatory postsynaptic current (EPSC) occurs during DSI of the temporally overlapping inhibitory postsynaptic current. With high levels of calcium chelators in the pipette solution, or bath application of bicuculline, EPSC enhancement is blocked, suggesting that it results from DSI and that the DSI process selectively affects GABAergic, but not glutamatergic, transmission. 3. The probability of synaptically evoked action potential firing is increased during DSI under current clamp. DSI could influence other excitatory phenomena as well.
Differentiation of the human monocytic cell line U937 results in an upregulation of the calcium release‐activated current, ICRAC.
Authors: Floto R A , Mahaut-Smith M P , Allen J M , Somasundaram B
Published: December 2014

1. Single cell fura‐2 fluorescence measurements and whole‐cell patch clamp recordings were used to investigate the effects of macrophage‐like differentiation, induced by dibutyryl cAMP (dbcAMP), on Ca2+ influx triggered by Ca2+ store depletion in the human monocytic cell line, U937. 2. In differentiated cells, the rise in intracellular [Ca2+] following store depletion by thapsigargin (TG) in nominally Ca(2+)‐free solution was 94% greater and the [Ca2+]i rise on subsequent re‐addition of external Ca2+ (2 mM) was 292% greater than in undifferentiated cells. 3. Under conditions where [Ca2+]i was buffered by BAPTA, TG‐induced store depletion failed to activate a detectable inward Ca2+ current in undifferentiated U937 cells. Under identical conditions, store depletion of differentiated U937 cells generated an inwardly rectifying Ca(2+)‐selective current which showed no reversal from ‐140 to +30 mV and was blocked by 1 microM external La3+; characteristics of the calcium release‐activated Ca2+ current (ICRAC) identified in other cells. 4. We conclude that U937 cells show a differentiation‐dependent upregulation of a store‐mediated Ca2+ entry pathway, identified as ICRAC, which is not correlated with the small associated increase in the size of TG‐sensitive Ca2+ pools.
Dopamine (D2) receptor regulation of intracellular calcium and membrane capacitance changes in rat melanotrophs.
Authors: Lee A K
Published: December 2014

1. Indo‐1 microfluorimetry and patch clamp techniques were used to study the decrease in cytosolic [Ca2+] ([Ca2+]i) caused by dopamine (D2) receptor activation and the calcium dependence of membrane capacitance changes in single rat melanotrophs. 2. [Ca2+]i decreased when extracellular calcium was removed or when the calcium channel blockers nickel (2 mM) or cadmium (100 microM) were applied by bath perfusion. 3. Quinpirole, a dopamine (D2) receptor agonist, reduced [Ca2+]i by 55 +/‐ 9 nM and hyperpolarized membrane potential by 29 +/‐ 9 mV simultaneously. 4. Quinpirole‐induced [Ca2+]i decrease required deactivation of voltage‐dependent calcium channels. Voltage clamping the membrane potential at ‐25 mV prevented the quinpirole‐induced [Ca2+]i decrease. Nickel (2 mM) reduced [Ca2+]i without hyperpolarization and precluded additional [Ca2+]i decrease by quinpirole. 5. Membrane capacitance measurement of secretion rates in cells dialysed with buffered calcium solutions showed that secretion began at approximately 400 nM Cai2+. 6. Melanotrophs have IP3‐sensitive calcium stores, but no caffeine‐sensitive calcium stores. Calcium released from IP3‐sensitive calcium stores also stimulated secretion. 7. Secretion in melanotrophs is modulated by protein kinase activators. cAMP (200 microM) enhanced secretion at [Ca2+]i > 1000 nM. Phorbol myristate acetate (PMA; 200 nM) enhanced secretion at [Ca2+]i < 400 nM, but not in the absence of calcium. 8. Dopamine receptor activation can reduce secretion by reducing the calcium influx through calcium channels with hyperpolarization of the membrane potential. However downregulation of either cAMP or protein kinase C activity may also contribute to the decrease in secretion.
Nitric oxide and exercise in the horse.
Authors: Mills P C , Marlin D J , Demoncheaux E , Scott C , Casas I , Smith N C , Higenbottam T
Published: December 2014

1. The effects of exercise on the production rate of nitric oxide (NO) in exhaled air (VNO) and the effects of inhaled NO (80 p.p.m.) on cardiovascular and respiratory parameters were investigated in five Throughbred horses. 2. The concentration of NO ([NO]) in exhaled air collected from within the nasal opening was lower when collected at a high flow rate of 80 l min‐1 than at a low flow rate of 20 l min‐1: when trotting at 3.7 m s‐1 the values were 0.78 +/‐ 0.15 and 1.23 +/‐ 9.14 p.p.b., respectively, and when cantering at 9 m s‐1 the values were 1.69 +/‐ 0.31 and 2.25 +/‐ 0.32 p.p.b., respectively. 3. Nebulized methoxamine (40 mg ml‐1 for 60 s), an alpha 1‐adrenergic agonist, further reduced [NO] during the 9 m s‐1 canter to 1.05 +/‐ 0.14 and 1.99 +/‐ 0.41 p.p.b. when collected at 80 and 20 l min‐1, respectively, and induced cyclical changes in the breathing pattern. 4. Exercise induced a linear increase in VNO with work intensity to a maximum (428.1 +/‐ 31.6 pmol min‐1 kg‐1) which coincided with the maximal oxygen uptake for the horses (138.3 +/‐ 11.7 ml min‐1 kg‐1), although a further increase in VNO (779.3 +/‐ 38.4 pmol min‐1 kg‐1) occurred immediately after exercise. The changes in VNO correlated well with the tidal volume (r = 0.968; P < 0.01) and the haematocrit (r = 0.855; P < 0.01). 5. In the first 2 min of high intensity exercise, inhaled NO (80 p.p.m.) significantly (P < 0.05) reduced the pulmonary artery pressure: during the first minute, pulmonary artery pressure was 83.1 +/‐ 7.6 mmHg compared with a control value of 94.4 +/‐ 6.3 mmHg, and during the second minute, 84.2 +/‐ 7.1 mmHg compared with a control value of 98.4 +/‐ 4.7 mmHg. There were no other significant changes in cardiovascular or respiratory indices, including cardiac output, measured during exercise between control and inhaled NO tests. 6. The results show that exhaled NO is released from the airways of the horse and may contribute to the regulation of pulmonary vascular tone during exercise.
Ca2+ mobilization and interlayer signal transfer in the heterocellular bilayered epithelium of the rabbit ciliary body.
Authors: Schütte M , Wolosin J M
Published: December 2014

1. ‘Ratiometric’ fura‐2 methodology in slice preparations and ‘intensitometric’ fluo‐3 measurements of confocal images were used to simultaneously monitor Ca2+ mobilization in the two distinct, apically joined cell layers which constitute the ciliary body epithelium (CBE): the non‐pigmented (NPE) and pigmented (PE) epithelia. 2. Both methods yielded comparable results regarding Ca2+ responses in the syncytium upon stimulation with adrenergic and cholinergic agonists. 3. The alpha 1‐adrenoceptor agonist phenylephrine elicited a moderate [Ca2+]i increase in the PE, whereas NPE [Ca2+]i remained unchanged or exhibited a slight diminution. 4. In combination with carbachol, the alpha 2‐adrenoceptor agonist brimonidine elicited large Ca2+ increases (> 10‐fold) in both the NPE and PE cell layers, even though previous studies indicated the absence of an alpha 2‐adrenergic effect on [Ca2+]i in the PE. The onset, as well as the peak of the Ca2+ responses in PE cells frequently exhibited a small delay with respect to adjacent NPE cells. No such time difference was observed between adjacent NPE cells. 5. Pre‐incubation of the ciliary body in Ca(2+)‐free solution under conditions known to elicit overt NPE‐PE separation abolished the alpha 2‐adrenocholinergic response in the PE. 6. Addition of heptanol to the perfusate, to block gap‐junctional communication, caused a small [Ca2+]i decrease in the NPE and a slight increase in PE[Ca2+]i. Subsequently, the Ca2+ mobilization in the Pe in response to the brimonidine and carbachol combination was either blocked or showed a substantial delay. The Ca2+ mobilization in the NPE, in contrast, remained unchanged. 7. We conclude that the heterocellular syncytium exhibits rectificatory behaviour with respect to Ca2+ mobilization; responses originating within the NPE are easily transferred to the PE, while the reverse does not occur.
Conducted depolarization in arteriole networks of the guinea‐pig small intestine: effect of branching of signal dissipation.
Authors: Segal S S , Neild T O
Published: December 2014

1. Blood flow control requires co‐ordinated activity among many branches of arteriole networks, which may be achieved by conduction of membrane potential changes between arteriolar smooth muscle cells and endothelial cells. 2. We investigated the effect of branching upon the passive conduction of electrical signals through the syncytium of electrically coupled cells in arteriole networks (n = 12) prepared from the guinea‐pig submucosa. To describe the effect of branching on cable properties, the expansion parameter B was calculated (B = 1 for an unbranched cable; B > 1 with branching) for a point in each arteriole network based on anatomy. 3. An estimate of B(B') was also obtained by measuring the spread of depolarization caused by a high‐K+ stimulus applied to one region. Membrane potential (‐74 +/‐ 4 mV (+/‐ S.D.) at rest) was recorded from smooth muscle cells (verified with intracellular dye labelling). A micropipette containing 120 mM KCl was positioned at 150 micron increments along an arteriole (width, 50‐75 microns) up to approximately 1.2 mm from a stationary recording site, producing stable depolarization which decreased as separation distance increased. The dissipation of depolarization with separation was greater when recording near branch origins rather than continuous segments. 4. B ranged in value from 0.99 to 2.28. In any one experiment, values of B and B' were correlated (correlation coefficient, r = 0.71; P < 0.05), but B' was consistently greater than B, and we discuss methodological factors which could lead to erroneously high values for B'. 5. For pooled electrophysiological data, depolarization decayed to 37% (1/e) of initial values in approximately 700 microns, consistent with B > 1. In contrast, the conduction of vasoconstriction and vasodilatation exceeds 2 mm in arteriole networks in previous studies. To explain this discrepancy, we suggest that active electrical events in cells of the arteriole wall augment passive electrical conduction during blood flow control.
Quantal properties of spontaneous EPSCs in neurones of the guinea‐pig dorsal lateral geniculate nucleus.
Authors: Paulsen O , Heggelund P
Published: December 2014

1. Spontaneous non‐NMDA glutamate receptor‐mediated EPSCs were recorded with the whole‐cell patch‐clamp technique from twenty‐six neurones in the dorsal lateral geniculate nucleus in thalamic slices from guinea‐pig. 2. Amplitude distributions of the EPSCs were skewed towards larger values. The skewness could be accounted for by multiquantal properties. The multiquantal properties were most clearly demonstrated in four cells that had prominent peaks in the amplitude distribution, and peak separation approximately corresponding to the modal value. The amplitude distribution for all cells could be adequately fitted by a quantal model consisting of a sum of Gaussians with means equal to integer multiples of a quantal unit. The variance of each Gaussian was equal to the sum of the noise variance of the recordings and an additional non‐negative variance which increased linearly with the number of the Gaussian in the series. The estimated mean quantal size was 152 +/‐ 37 pS. The estimated mean quantal coefficient of variation was 15%. Addition of tetrodotoxin did not significantly change any of the quantal parameters (n = 5). 3. The waveform of the EPSCs was similar for small and large events, and similar to that of events evoked by stimulation of retinal input fibres. There was a positive correlation between peak amplitude and rise time. This is the opposite of that expected if differences in electrotonic distances were to explain differences in amplitude. 4. The spontaneous EPSCs occurred randomly at an average frequency of 3.1 Hz. The events with amplitudes approximately equal to multiples of the quantal size were, in most cells, more numerous than could be accounted for by coincidence of randomly occurring events of quantal size. 5. The results indicate that spontaneous EPSCs can reflect more than a single quantum, and suggest that quantal events may be coupled due to action potential‐independent near‐synchronous multiquantal release of transmitter.
The action of cytoplasmic calcium on the cGMP‐activated channel in salamander rod photoreceptors.
Authors: Sagoo M S , Lagnado L
Published: December 2014

1. Truncated salamander rod photoreceptors were internally perfused to investigate the action of cytoplasmic Ca2+ on cGMP‐activated channels in the outer segment. 2. Switching from 1 microM Ca2+ to 0 Ca2+ increased the cGMP‐activated current by a factor of 7.1 +/‐ 0.5 when measured in the first 60 s after the outer segment was opened to the bath, but only 2‐fold after 5 min or more. This was attributed to the loss from the outer segment of a soluble factor required for Ca2+ to inhibit the cGMP‐activated channel. 3. Short exposures to 0 Ca2+ caused an irreversible increase in the cGMP‐activated current measured in 1 microM Ca2+, indicating that lowering [Ca2+] accelerated the loss of the channel inhibitor from the outer segment. 4. Channel activation occurred with a half‐time of 6.7 s on switching to 0 Ca2+. Replacing 1 microM Ca2+ inhibited the current again with a half‐time of 11.0 s. 5. The inhibition of the cGMP‐activated current by Ca2+ could be described by a Hill curve with half‐maximal suppression at 55 +/‐ 13 nM Ca2+ and a Hill coefficient of 1.4 +/‐ 0.4. 6. Addition of calmodulin (1 microM), or the calmodulin inhibitors mastoparan and calmidazolium (5 microM), did not alter the action of Ca2+ on the cGMP‐activated current. 7. The increased affinity of the cGMP‐activated channels in response to a fall in [Ca2+] has the magnitude, speed and Ca2+ dependence to suggest that it will promote recovery of the cGMP‐activated current in response to the light‐induced fall in [Ca2+] that normally occurs inside the outer segment.