bluuebunny/crossref_metadata_2025 huggingface dataset:
Responsiveness of cat area 17 after monocular inactivation: limitation of topographic plasticity in adult cortex.
Authors: Rosa M G , Schmid L M , Calford M B
Published: December 2014

1. Recordings were made from neurones in the splenial sulcus of normal adult cats and adult cats which had one eye inactivated by enucleation or photocoagulation of the optic disc. Two visually responsive regions were observed, corresponding to the peripheral representation of visual area 1 (V1) and the splenial visual area. In normal animals, responses to the ipsilateral eye in V1 were restricted to the medial half of the splenial sulcus, up to 45‐50 deg eccentricity. Thus, by inactivating the eye contralateral to the experimental hemisphere, we created a region in V1, 1‐2 mm wide, that lacked normal inputs. 2. In contrast to results from previous experiments where lesions were placed in the central retina, neurones in the deprived peripheral representation remained unresponsive to light stimuli for up to 12 h after deactivation of the contralateral eye. 3. In animals that were allowed to recover from the monocular deactivation for periods of 2 days to 16 months, there was rearrangement of the retinotopic maps. Receptive fields in regions of cortex that normally represented the monocular crescent were displaced to the temporal border of the binocular field of vision. However, most neurones in the deprived peripheral representation remained unresponsive to visual stimuli even more than 1 year after treatment. This is also in marked contrast with the extensive reorganization that is observed in the central representation of V1 after restricted retinal lesions. Analysis of the cortical magnification factor demonstrates that the change in visual topography is local, and does not involve an overall centro‐peripheral shift of the retinotopic map. 4. Among the neurones that did show displaced receptive fields, the response properties were clearly abnormal. They showed a notable lack of spontaneous activity, low firing rates and rapid habituation to repeated stimulation. 5. The low potential for reorganization of the monocular sector of V1 demonstrates that the capacity for plasticity of mature sensory representations varies with location in cortex. Even relatively small pieces of cortex, such as the monocular crescent representations, may not reorganize completely if certain conditions are not met. These results suggest the existence of natural boundaries that may limit the process of reorganization of sensory representations.
The effect of partial denervation of developing rats fast muscles on their motor unit properties.
Authors: Tyc F , Vrbová G
Published: December 2014

1. The effects of partial denervation on motor units of the fast twitch extensor digitorum longus (EDL) muscle of the rat were studied. 2. Partial denervation was performed by surgically removing 2‐4 mm of the L4 ventral ramus in 3‐ and 18‐day‐old Wistar rats. Two to three months later, EMG activity, contractile properties and muscle fibre types were analysed. 3. After partial denervation the EDL muscle became significantly more active, particularly in the animals operated on at 3 days. The amount of activity during periods of rest was up to 4 times that of the control EDL muscle. 4. The maximum tetanic tension developed by the EDL muscles 2 months after partial denervation at 3 days was only 11 +/‐ 1.01% (S.E.M., n = 18) of the control. In animals operated on at 18 days this value was larger, i.e. 44 +/‐ 3.46% (S.E.M., n = 12). The low force output of animals operated on at 3 days was also reflected in the low mean motor unit (MU) force output which was only 69 +/‐ 5.82% (S.E.M., n = 17) of the contralateral control muscle. In contrast the force generated by MUs of rats operated on at 18 days was larger than that of control muscles, i.e. 151 +/‐ 13.05% (S.E.M., n = 11). The number of MUs was 6 +/‐ 0.32 (S.E.M., n = 19) in rats operated on at 3 days and 12 +/‐ 0.83 (S.E.M., n = 14) in rats operated on at 18 days. 5. The speed of contraction decreased and the resistance to fatigue increased. These changes were greater in animals operated on at 3 days. The proportion of muscle fibres reacting with antibody against slow myosin showed a significant increase, especially in the group of animals operated on at 3 days.
Red‐green opponent channel mediation of control of human ocular accommodation.
Authors: Kotulak J C , Morse S E , Billock V A
Published: December 2014

1. It has been hypothesized, but not verified empirically, that the control of human ocular accommodation is mediated by either the red‐green or yellow‐blue colour channels. Our goal was to determine experimentally whether the red‐green channel by itself could influence the accommodative response. 2. To find out, we isolated the red‐green channel through chromatic bandpass filtering and measured accommodation under dynamic and static conditions. The effect of this filtering was to modulate the red‐green channel without disturbing either the yellow‐blue or luminance channels. 3. Accommodative gain (ratio of response to stimulus amplitude) declined monotonically with decreasing bandwidth under dynamic conditions. Because the outputs of both the luminance and yellow‐blue colour channels did not vary with bandwidth, the only explanation is that the red‐green opponent process was responsible for the effect. 4. Under static conditions, however, accommodation was independent of bandwidth. This may be attributable to the decreased sensitivity to chromatic contrast that occurs at low temporal frequencies.
Expression and distribution of sodium channels in short‐ and long‐term denervated rodent skeletal muscles.
Authors: Lupa M T , Krzemien D M , Schaller K L , Caldwell J H
Published: December 2014

1. Loose‐patch voltage‐clamp recordings were made from rat and mouse skeletal muscle fibres denervated for up to 6 weeks. Innervated muscles possessed a Na+ current density of 107 +/‐ 3.3 mA cm‐2 in endplate membrane, and 6.3 +/‐ 0.6 mA cm‐2 in extrajunctional membrane. This high concentration of Na+ channels at the endplate was gradually reduced following denervation. After 6 weeks of denervation, the endplate Na+ channel concentration was reduced by 40‐50%, and the density of Na+ channels in extrajunctional membrane was increased by about 30%. 2. The tetrodotoxin (TTX)‐resistant form of the Na+ channel appeared after 3 days of denervation and comprised approximately 43% of the endplate Na+ channels 5‐6 days after denervation. Subsequently, TTX‐resistant Na+ channels were reduced in density to approximately 25% of the postjunctional Na+ channels and remained at this level up to 6 weeks after denervation. 3. RNase protection analysis showed that mRNA encoding the TTX‐resistant Na+ channel was virtually absent in innervated muscle, rose > 50‐fold after 3 days of denervation, then decreased by 95% 6 weeks after denervation. The density of TTX‐resistant Na+ channels correlated qualitatively with changes in mRNA levels. 4. These results suggest that the density of Na+ channels at neuromuscular junctions is maintained by two mechanisms, one influenced by the nerve terminal and the other independent of innervation.
Regulation of granule size in human and horse eosinophils by number of fusion events among unit granules.
Authors: Hartmann J , Scepek S , Lindau M
Published: December 2014

1. We have investigated the granule size distributions in human and horse eosinophils by time‐resolved patch‐clamp capacitance measurements. 2. During exocytosis of single granules the electrical capacitance of the plasma membrane increases in discrete steps. The steps in horse cells are about six times larger than those in human cells in accordance with the difference in granule size. 3. In both species a multimodal capacitance step size distribution is observed with a first peak at 6‐7 fF corresponding to granules with a diameter of about 450‐500 nm and a surface area of about 0.7 microns2, which we call the unit granule. The other peaks in the distributions correspond to multiples of the surface area of these units. 4. These results show that the larger granules are formed by fusion of several unit granules and the final size of mature granules is determined by the number of units allowed to fuse with each other. Whereas in human eosinophils most granules consist of one or two units, most granules of horse eosinophils are formed by fusion of seven to fifteen units. 5. The intracellular fusion events associated with vesicular traffic are believed to occur constitutively. In contrast, our results indicate that a cellular mechanism exists which regulates the size of the mature granules by determining the number of units allowed to fuse with each other. In view of our recent report that granule‐granule fusion can be activated by GTP gamma S, this regulation may possibly involve GTP‐binding proteins.
Organization of neuronal systems mediating presynaptic inhibition of group II muscle afferents in the cat.
Authors: Riddell J S , Jankowska E , Huber J
Published: December 2014

1. The organization of neuronal systems mediating presynaptic control of transmission from group II muscle afferent fibres has been investigated by comparing the sources of presynaptic inhibition of fibres terminating in different segments of the spinal cord: fibres of the semitendinosus and lateral gastrocnemius muscle nerves terminating in the sacral segments and of the tibialis anterior and extensor digitorum longus muscle nerves terminating in the midlumbar segments. 2. Two measures of presynaptic inhibition were used: depolarization of the terminals of group II fibres (detected as changes in the excitability of single fibres to electrical stimuli) and a decrease in the effectiveness of their synaptic actions (detected as a decrease in the amplitude of monosynaptic field potentials evoked by group II muscle afferents). 3. Group II muscle afferents strongly depolarized all of the group II afferent fibres, while group I muscle afferents contributed to the depolarization of only a few. The majority of fibres were as effectively depolarized by cutaneous afferents as by the most effective muscle afferents. However, the effectiveness with which afferents of different nerves depolarized group II muscle afferent fibres in the sacral and midlumbar segments differed. The most effective afferents were those of nerves that provide the main input to dorsal horn interneurones in the same region of the spinal cord. The sources of depolarization of flexor and extensor fibres terminating in the same (sacral) segments were very similar. 4. The amplitudes of field potentials evoked by group II afferents were depressed by the same types of afferent as produced depolarization of group II afferent fibres. There was also a strong correlation between the effectiveness with which afferents of a given nerve induced depolarization of single fibres and depression of field potentials in the same segments. Since group II field potentials were depressed to a greater extent (by up to 90%) than group I field potentials (by no more than 20%) concurrently recorded in the intermediate zone of midlumbar segments, it appears that transmission from group II muscle afferents may be more strongly affected by presynaptic inhibition than that from group I muscle afferents. 5. The results suggest that the interneuronal systems responsible for the presynaptic control of transmission from group II muscle afferents have topographically restricted actions and an organization appropriate to a system of negative feedback control.
Vascular effects of parathyroid hormone and parathyroid hormone‐related protein in the split hydronephrotic rat kidney.
Authors: Endlich K , Massfelder T , Helwig J J , Steinhausen M
Published: December 2014

1. The effects of locally applied parathyroid hormone‐related protein (PTHRP), a putative autocrine/paracrine hormone, on vascular diameters and glomerular blood flow (GBF) in the split hydronephrotic rat kidney were studied. As PTHRP interacts with parathyroid hormone (PTH) receptors in all tissues tested so far, the effects of PTHRP were compared with those of PTH. 2. Preglomerular vessels dilated in a concentration‐ and time‐dependent manner that was almost identical for PTH and PTHRP. A significant preglomerular vasodilation (5‐17%) occurred at a threshold concentration of 10(‐10) mol l‐1 PTH or PTHRP, which raised GBF by 20 +/‐ 2 and 31 +/‐ 4%, respectively (means +/‐ S.E.M., n = 6). PTH or PTHRP (10(‐7) mol l‐1) increased preglomerular diameters (11‐36%) and GBF (60 +/‐ 10 and 70 +/‐ 8%, respectively) to near maximum. The most prominent dilatation was located at the interlobular artery and at the proximal afferent arteriole. 3. Efferent arterioles were not affected by either PTH or PTHRP. 4. Estimated concentrations of half‐maximal response (EC50) for preglomerular vasodilatation and GBF increase were in the nanomolar to subnanomolar range. 5. After inhibition of angiotensin I‐converting enzyme by 2 x 10(‐6) mol kg‐1 quinapril I.V. (n = 6), 10(‐8) mol l‐1 PTHRP dilated preglomerular vessels and efferent arterioles (9 +/‐ 1% proximal and 6 +/‐ 1% distal). 6. We conclude that the renal vasculature of the hydronephrotic kidney is highly sensitive to vasodilatation by PTH and PTHRP, which, in addition, may constrict efferent arterioles by stimulating renin release.(ABSTRACT TRUNCATED AT 250 WORDS)
Calcium‐activated potassium channels in human platelets.
Authors: Mahaut-Smith M P
Published: December 2014

1. The effect of intracellular [Ca2+] ([Ca2+]i) on human platelet ion channels was studied using the nystatin whole‐cell patch clamp recording technique. 2. Ionomycin‐induced increases in [Ca2+]i rapidly activated a voltage‐independent K(+)‐selective channel with a slope conductance of 30 pS in 154 mM K+ saline. The single‐channel conductance decreased in proportion to the square root of the external K+ concentration such that the estimated conductance in 5 mM K+ was approximately 5 pS. 3. The peak current under conditions expected to increase [Ca2+]i to micromolar levels indicated that each platelet possesses a small number (5‐7) of 30 pS Ca(2+)‐dependent K+ channels (KCa channels). 4. Spontaneous [Ca2+]i spiking was observed in many patch‐clamped platelets using fura‐2 fluorescence measurements. Each Ca2+ spike triggered up to five KCa channels at any one time. KCa channels were not active at resting levels of [Ca2+]i. 5. The results suggest that platelet KCa channels are not active under resting conditions but may have an important role in determining the membrane potential during Ca2+ signalling.
Cholinergic and electrical synapses between synergistic spinal motoneurones in the Xenopus laevis embryo.
Authors: Perrins R , Roberts A
Published: December 2014

1. To investigate central motoneurone synapses within the spinal cord of a simple vertebrate, the Xenopus embryo, simultaneous intracellular recordings were made from fifty‐five pairs of spinal motoneurones. 2. Chemical synapses were found between seventeen out of thirty‐five pairs on the same side of the spinal cord. Current‐evoked spikes in the presynaptic neurone led to fast depolarizing postsynaptic potentials (PSPs) in the postsynaptic neurone at latencies of 0.5‐1.5 ms. The PSPs had an average amplitude of 7 mV, a rise time of 8 ms and a half‐fall time of 18 ms. 3. The presynaptic motoneurone was always the more rostral of the pair. No excitatory connections were found which crossed the cord. The fast PSPs were blocked by 10 microM mecamylamine but not by 1 mM kynurenic acid, so were mediated by nicotinic acetylcholine receptors (nAChRs). These are the first unitary excitatory postsynaptic potentials (EPSPs) mediated by nAChRs recorded intracellularly within the vertebrate central nervous system. 4. Bidirectional electrical synapses were found between five pairs of motoneurones. All these pairs were on the same side of the spinal cord and less than 70 microns apart. Each neurone responded in a graded manner to either hyperpolarizing or depolarizing current injected into the other. 5. Since motoneurones are rhythmically active during swimming, both chemical and electrical synapses will add to the fast on‐cycle excitation underlying spiking activity in other motoneurones. This may increase the reliability and local synchrony of synergistic motoneurone firing during locomotion.
Effect of intra‐articular hyaluronan on pressure‐flow relation across synovium in anaesthetized rabbits.
Authors: McDonald J N , Levick J R
Published: December 2014

1. Hyaluronan is the major polysaccharide of synovial fluid, responsible for its high viscosity. The effect of hyaluronan on fluid transport across the synovial lining of the joint was investigated. Rate of fluid absorption from the joint cavity (Qs) was measured at intra‐articular pressures (Pj) of up to 24 cmH2O in knees of anaesthetized rabbits, in the presence or absence of hyaluronan in intra‐articular infusates. 2. Viscometry studies in vitro showed that the commercial hyaluronan used had a molecular weight of 549,000‐774,000, a radius of gyration of 48‐99 nm and a critical concentration for molecular overlap of 1.3 g l‐1. 3. With intra‐articular Krebs solution (control) or subnormal, subcritical concentrations of hyaluronan (0.5 g l‐1), flow increased with pressure. Hyaluronan reduced the fluid escape rate by reducing slope dQs/dPj by 32‐64% relative to Krebs solution. 4. At normal to high hyaluronan concentrations (3‐6 g l‐1) and low pressures, hyaluronan again reduced slope dQs/dPj, by 39‐64%. The reduction in slope was slight, however, when compared with the reduction in bulk fluidity (1/relative viscosity). Fluidity at high shear rates was reduced to 6% of control values by 6 g l‐1 hyaluronan. The effect on slope did not correlate significantly with the effect on fluidity. 5. At pressures above approximately 12 cmH2O, 3‐6 g l‐1 hyaluronan altered the shape of the pressure‐flow relation: a flow plateau developed. In some joints raising pressure even reduced trans‐synovial flow slightly. The pressure required to drive unit trans‐synovial flow (an index of outflow resistance) increased 2.5‐fold between 5 and 25 cmH2O in the presence of hyaluronan. By contrast, in the absence of hyaluronan the outflow resistance fell as pressure was raised. 6. It is suggested that the increasing resistance to flow in the presence of hyaluronan may be caused by partial molecular sieving of hyaluronan by the small porosities of the synovial interstitial matrix, leading to accumulation of a resistive filter cake of hyaluronan chains at the tissue‐cavity interface. Since hyaluronan impedes fluid escape when pressure is raised, it may serve to preserve synovial fluid volume in vivo, e.g. during sustained joint flexion.
Inhibition of the interaction of G protein G(o) with calcium channels by the calcium channel beta‐subunit in rat neurones.
Authors: Campbell V , Berrow N S , Fitzgerald E M , Brickley K , Dolphin A C
Published: December 2014

1. The beta‐subunit has marked effects on the biophysical and pharmacological properties of voltage‐dependent calcium channels. In the present study we examined the ability of the GABAB agonist (‐) ‐baclofen to inhibit calcium channel currents in cultured rat dorsal root ganglion neurones following depletion of beta‐subunit immunoreactivity, 108‐116 h after microinjection of a beta‐subunit antisense oligonucleotide. 2.We observed that, although the calcium channel current was markedly reduced in amplitude following beta‐subunit depletion, the residual current (comprising both N‐ and L‐type calcium channel currents) showed an enhanced response to application of (‐) ‐baclofen. Therefore, it is possible that there is normally competition between activated G protein G(o) and the calcium channel beta‐subunit for binding to the calcium channel alpha 1‐subunit; and this competition shifts in favour of the binding of activated G(o) following depletion of the beta‐subunit, resulting in increased inhibition. 3. This hypothesis is supported by evidence that an antibody against the calcium channel beta‐subunit completely abolishes stimulation of the GTPase activity of G(o) by the dihydropyridine agonist S‐(‐) ‐Bay K 8644 in brain membranes. This stimulation of GTPase is thought to result from an interaction of G(o) alpha‐subunit (G alpha o) with its calcium channel effector which may operate as a GTPase‐activating protein. 4. These data suggest that the calcium channel beta‐subunit when complexed with the beta 1‐subunit normally inhibits its association with activated G(o). It may function as a GTPase‐activating protein to reduce the ability of activated G(o) to associate with the calcium channel, and thus limit the efficacy of agonists such as (‐) ‐baclofen.
Hypoxia‐induced brain angiogenesis in the adult rat.
Authors: Harik S I , Hritz M A , LaManna J C
Published: December 2014

1. Prolonged hypoxia increases the brain vascularity. Here we studied the protein and deoxyribonucleic acid (DNA) content of isolated cerebral microvessels in hypoxic and control rats. 2. Adult male Wistar rats that were subjected to hypobaric hypoxia at 50.5 kPa (0.5 atm) for periods of 1, 2, or 3 weeks and normoxic littermate controls were used. Cerebral microvessels were harvested by bulk isolation from the pooled cortical mantles of three to six rats in each group. The isolated microvessels were assayed for their protein and DNA content. 3. Microvessel protein yield increased from 0.31 to 0.45 mg of microvessel protein (g wet wt)‐1 of cerebral cortex after 1 week of hypoxia, but did not increase further during up to 3 weeks of hypoxia. In contrast, microvessel DNA yield did not increase during the first week of hypoxia, but increased significantly after 2 weeks of hypoxia and continued to increase up to 56 micrograms of microvessel DNA (g cerebral cortex)‐1 at 3 weeks of hypoxia (normoxic mean was 32 micrograms DNA g‐1). 4. The cell size index (mg protein:mg DNA) of isolated cerebral microvessels increased after 1 week of hypoxia, suggesting microvascular hypertrophy, but returned to control by the second week of hypoxia and decreased to below control levels by the third week of hypoxia, suggesting microvascular hyperplasia. These results indicate that the increased vascularity of the brain in hypobaric hypoxia progresses from an early phase of microvascular hypertrophy to later microvascular hyperplasia.
Opioid depression of respiration in neonatal rats.
Authors: Greer J J , Carter J E , al-Zubaidy Z
Published: December 2014

1. The effects of opioid receptor agonists and antagonists on the breathing pattern of neonatal rats were studied. Three experimental approaches were taken. In the first approach, the effects of opioid agonists and antagonists on the spontaneous respiratory neural activity generated by brainstem‐spinal cords isolated from neonatal rats aged 0‐4 days postnatal (P0‐4) maintained in vitro were studied. Secondly, similar studies were performed utilizing medullary slice preparations consisting of respiratory rhythm‐generating regions (pre‐Bötzinger complex). Thirdly, whole‐body plethysmographic recordings were obtained from unanaesthetized neonatal (P0‐18) rats before and after I.P. administration of opioid‐receptor agonists and antagonists. 2. The mu‐receptor agonists morphiceptin and DAGO (Tyr‐D‐Ala‐Gly‐[NMePhe]‐Gly‐ol), when added either to the solutions bathing the brainstems of neonatal rat brainstem‐spinal cord preparations or bathing the medullary slice preparations, resulted in a naloxone‐reversible, dose‐dependent decrease in the frequency of respiratory rhythmic discharge. 3. The respiratory burst frequency and amplitude in vitro were unaffected by the addition of the delta‐opioid receptor agonist DPDPE ([D‐pen2,5]‐enkephalin) and the kappa‐opioid receptor agonist U50488 (trans‐[+]‐3,4‐dichloro‐N‐methyl‐N‐(2‐[1‐ pyrrolidinyl]cyclohexyl) benzene‐acetamide) or the opioid receptor antagonist naloxone. 4. Intraperitoneal administration of the mu‐opioid receptor agonist fentanyl resulted in a naloxone‐reversible, dose‐dependent decrease in the frequency and amplitude of breathing of unanaesthetized neonatal rats (P0‐P10). I.P. administration of the delta‐opioid receptor agonist DPDPE did not affect breathing of neonatal rats until the second week postnatally. 5. We conclude that opioids suppress the frequency of neonatal rat respiration by acting via mu‐opioid receptors located within regions of the ventral medulla containing respiratory rhythm‐generating centres (the pre‐Bötzinger complex). delta‐Opioid receptor activation does not affect breathing in neonatal rats until approximately the second week postnatally.
Ca(2+)‐induced Ca2+ release and its activation in response to a single action potential in rabbit otic ganglion cells.
Authors: Yoshizaki K , Hoshino T , Sato M , Koyano H , Nohmi M , Hua S Y , Kuba K
Published: December 2014

1. Ryanodine‐sensitive intracellular Ca2+ release activated by Ca2+ entry was studied with fura‐2 fluorescence and intracellular voltage recording techniques in rabbit otic ganglion cells. 2. The removal of extracellular Ca2+ reduced sustained, transient or oscillatory rises in intracellular Ca2+ ([Ca2+]i) induced at high extracellular K+ and abolished the [Ca2+]i oscillation in cultured neurones. 3. Ryanodine (10 microM) transiently increased [Ca2+]i and reduced the amplitude and rate of rise of the high‐K(+)‐induced rise in [Ca2+]i, while caffeine (5 mM) produced a few transient rises in [Ca2+]i in most cultured cells and [Ca2+]i oscillation only in one cell. 4. The two components of the slow after‐hyperpolarization (AHP) of an action potential in neurones of freshly isolated ganglia were dependent on extracellular Ca2+ and abolished by Ca2+ channel blockers, Cd2+ or Co2+. 5. The late component of AHP (LAHP), but not the initial component, in ‘fresh’ neurones increased in area with an increase in the preceding interval, was abolished by ryanodine (10 microM) and intracellularly injected EGTA, and mimicked by intracellular injection of Ca2+. 6. A ryanodine‐sensitive Ca(2+)‐induced Ca2+ release thus exists, operates in response to an action potential‐induced Ca2+ entry and underlies LAHP in rabbit otic ganglion cells.
Amino acid‐induced [Ca2+]i oscillations in single mouse pancreatic islets of Langerhans.
Authors: Martin F , Soria B
Published: December 2014

1. The effects of amino acids on cytosolic free calcium concentration ([Ca2+]i) were measured, using fura‐2 fluorescence imaging, in mouse pancreatic islets of Langerhans. 2. Slow [Ca2+]i oscillations appeared when isolated islets were incubated with a solution containing a mixture of amino acids and glucose at concentrations found in the plasma of fed animals. 3. In the presence of 11 mM glucose, alanine (5 mM) and arginine (10 mM) induced a transient rise in [Ca2+]i followed by an oscillatory pattern, while leucine (3 mM) and isoleucine (10 mM) triggered the appearance of slow [Ca2+]i oscillations. 4. Also in the presence of glucose (11 mM), tolbutamide (10 microM) increased the duration of the glucose‐induced [Ca2+]i oscillations. While tolbutamide (10 microM) did not modify the leucine‐induced slow oscillatory pattern, addition of diazoxide (10 microM) resulted in the gradual appearance of [Ca2+]i oscillations which resembled the glucose‐induced fast oscillations. 5. Like stimulatory glucose concentrations (11 mM), glyceraldehyde (10 mM) induced fast oscillations of [Ca2+]i. 6. Fluoroacetate (2 mM) transformed leucine‐induced slow [Ca2]i oscillations into fast [Ca2+]i oscillations. Iodoacetate (1 mM) completely inhibited any oscillatory pattern. 7. It is suggested that mitochondrially generated signals, derived from amino acid oxidative metabolism, acting in conjunction with glucose‐signalled messengers, are very effective at closing ATP‐dependent K+ channels (KATP+). 8. We propose that metabolic regulation of KATP+ channels is one of the mechanisms underlying the modulation of the oscillatory [Ca2+]i response to nutrient secretagogues.
beta‐adrenergic and cholinergic modulation of the inwardly rectifying K+ current in guinea‐pig ventricular myocytes.
Authors: Koumi S , Wasserstrom J A , Ten Eick R E
Published: December 2014

1. Whole‐cell patch‐clamp technique was used to study the beta‐adrenergic and cholinergic regulation of the inwardly rectifying K+ conductance (gK1) in isolated guinea‐pig ventricular myocytes. 2. In Cl(‐)‐free solutions or in the presence of 9‐anthracenecarboxylic acid or Co2+, bath‐applied isoprenaline (Iso) partially inhibited the steady‐state whole‐cell conductance (gss) calculated from the steady‐state current (Iss)‐voltage (Iss‐V) curve at membrane voltages (Vm) negative to the equilibrium potential for potassium (EK). Iss was also inhibited at Vm positive to EK when the extracellular [K+] was 20 mM. The Iso‐sensitive component of gss exhibited the characteristics of the inwardly rectifying K+ conductance (gK1). 3. The Iso‐induced inhibition of gK1 was reversible, concentration dependent, blocked by propranolol, mimicked by both forskolin and dibutyryl cAMP, and prevented by including a cAMP‐dependent protein kinase (PKA) inhibitor in the pipette solution. These findings suggest that PKA mediates the Iso‐induced inhibition of gK1. 4. The apparent dissociation constant (KD) for the concentration dependence of Iso‐induced inhibition was 0.035 microM and the Hill coefficient was approximately 1.0. A maximal Iso concentration (1 microM) inhibited gK1 by 40 +/‐ 4.1% (mean +/‐ S.E.M.; n = 13). 5. Bath application of acetylcholine (ACh, 0.1 microM or more) antagonized the Iso‐induced (1 microM) inhibition of gK1; [ACh] > 1.0 microM antagonized 88 +/‐ 2.1% (n = 10) of the inhibition. ACh increased the KD for Iso to inhibit Iso‐sensitive gK1 and also reduced the maximal Iso‐induced inhibition. 6. ACh‐induced antagonism could be abolished by pre‐incubating myocytes with pertussis toxin (PTX), suggesting that a muscarinic receptor‐coupled, PTX‐sensitive G protein, Gi, is involved. 7. ACh (10 microM) also antagonized approximately 70% of the dibutyryl cyclic AMP (1 mM)‐induced inhibition of gK1 (n = 3), suggesting that the ACh‐induced antagonism involves more than simply inhibiting the Iso‐mediated activation of adenylyl cyclase via the activated Gi. 8. Intracellularly applied okadaic acid (OkA, 1 microM) did not alter gK1 (control = 134 +/‐ 5.1 nS vs. OkA = 136 +/‐ 6.1 nS), but the Iso‐induced decrease in gK1 was less (P < 0.001) with OkA present (42.1 +/‐ 2.4 nS, n = 5) than when absent (54.0 +/‐ 2.2 nS, n = 10). However, ACh (10 microM) failed to antagonize Iso‐induced inhibition with OkA present, suggesting involvement of a protein phosphatase.
Phosphoarginine stimulation of Na(+)‐Ca2+ exchange in squid axons‐‐a new pathway for metabolic regulation?
Authors: DiPolo R , Beaugé L
Published: December 2014

1. [Na+]o‐dependent Ca2+ efflux (forward Na(+)‐Ca2+ exchange), [32P]ATP wash‐out curves and [ATP] were measured in internally dialysed squid giant axons at 17‐18 degrees C. 2. We found that dialysing squid axons without ATP and with [Ca2+]i around 1 microM the basal levels of the [Na+]o‐dependent Ca2+ efflux were significantly higher in the presence of N omega‐phosphoarginine (PA). Phosphocreatine, a related phosphagen, is without effect. 3. PA stimulation of the Na(+)‐Ca2+ exchange occurs in the complete absence of ATP (< 1 microM), being independent of, and additive to, the ATP‐stimulated [Na+]o‐dependent Ca2+ efflux. PA stimulation of [Na+]o‐dependent Ca2+ efflux is fully and rapidly reversible with a Km around 7.7 mM. Activation by saturating [PA] is equivalent in magnitude to that of ATP. 4. PA stimulation of Na(+)‐Ca2+ exchange is markedly dependent on intracellular Ca2+ and Mg2+ ions. Below 0.5 microM Ca2+i PA effect is negligible, becoming noticeable between 0.8 and 2 microM. In addition, Ca2+i considerably increases the rate at which PA activates the Na(+)‐Ca2+ exchange. Although there is no absolute requirement of the PA effect for Mg2+ ions, this divalent cation largely stimulates the PA effect. 5. This work demonstrates, for the first time, the presence in squid axons of a new form of metabolic regulation of the Na(+)‐Ca2+ exchange directly and solely related to PA and different from that of MgATP. This novel mechanism is likely to play a physiological role in Ca2+ extrusion through the Na(+)‐Ca2+ exchanger, particularly at micromolar [Ca2+]i.
Mechanisms of intracellular Mg2+ regulation affected by amiloride and ouabain in the guinea‐pig taenia caeci.
Authors: Nakayama S , Nomura H
Published: December 2014

1. The effects of amiloride and ouabain on the regulation of the intracellular, free Mg2+ concentration ([Mg2+]i) were investigated in the taenia isolated from the guinea‐pig caecum, using nuclear magnetic resonance (NMR) techniques. 2. [Mg2+]i were mainly estimated from the separation of the alpha‐ and beta‐ATP peaks observed in 31P NMR spectra. In normal (physiological) and nominally Ca(2+)‐free solutions, [Mg2+]i was approximately 0.3‐0.4 mM. Application of either amiloride or ouabain in Ca(2+)‐free solutions significantly increased [Mg2+]i, with only a small change in ATP content. Washout of the drugs reversed the changes in [Mg2+]i. 3. Changes in pHi were estimated from: (1) the chemical shift of phosphoethanolamine, and (2) solving two relational equations of pHi and [Mg2+]i obtained from the beta‐ and gamma‐ATP peaks. Both estimations revealed some intracellular alkalosis during application of these two drugs. After correction for pHi, a significant increase in [Mg2+]i was still obtained 150 min after application of either drug. 4. In the presence of amiloride, simultaneous removal of extracellular Mg2+ and Ca2+ significantly depleted intracellular Mg2+. This result suggests the presence of an amiloride‐insensitive (or less sensitive) pathway which passively transports Mg2+ across the plasma membrane. 5. The intracellular Rb+ concentration was monitored as an index of Na(+)‐K+ pump activity, using 87Rb NMR. In Ca(2+)‐free solutions containing 5 mM Rb+, the intracellular Rb+ concentration was hardly changed by amiloride, but was depleted by additional applications of ouabain. Wash‐out of ouabain restored the intracellular Rb+ in the presence of amiloride. 6. These results are consistent with the presence of Na(+)‐Mg2+ exchange as an effective Mg(2+)‐extruding mechanism in smooth muscle. Although many other factors may cause changes in [Mg2+]i, it seems likely that amiloride directly inhibits the Na(+)‐Mg2+ exchanger, whilst ouabain does so indirectly through reduction of the Na+ gradient across the plasma membrane.
Two distinct modes of Ca2+ signalling by ACh in rat pancreatic beta‐cells: concentration, glucose dependence and Ca2+ origin.
Authors: Yada T , Hamakawa N , Yaekura K
Published: December 2014

1. Calcium signalling by acetylcholine (ACh) in single rat pancreatic beta‐cells was studied. The cytosolic free Ca2+ concentration ([Ca2+]i) was measured by dual‐wavelength fura‐2 microfluorometry. 2. In the presence of basal glucose (2.8 mM), 10(‐6) to 10(‐4) M ACh (high ACh) transiently increased [Ca2+]i. The [Ca2+]i response to 10(‐5) M ACh was little altered under Ca(2+)‐free conditions. Brief pulses of 10(‐5) M ACh evoked successive [Ca2+]i responses, which were progressively inhibited by 0.2‐0.5 microM thapsigargin, a specific inhibitor of the endoplasmic reticulum (ER) Ca2+ pump. 3. Elevation of glucose to 8.3 mM, a concentration which stimulates insulin release, increased [Ca2+]i to an initial peak followed by a sustained, moderate elevation. Addition of 10(‐8) to 10(‐7) M ACh (low ACh) evoked a further increase in [Ca2+]i. The [Ca2+]i response to 10(‐7) M ACh was completely inhibited under Ca(2+)‐free conditions by 1 microM nitrendipine, a blocker of L‐type Ca2+ channels, and by 100 microM diazoxide, an opener of ATP‐sensitive K+ channels. 4. In the presence of 8.3 mM glucose, [Ca2+]i responses to 10(‐5) M ACh were reduced but not abolished by Ca(2+)‐free conditions, nitrendipine and diazoxide. Successive [Ca2+]i transients induced by 10(‐5) M ACh pulses in the presence of nitrendipine were progressively inhibited by thapsigargin. 5. The results revealed two distinct modes of Ca2+ signalling: low ACh increases [Ca2+]i by stimulating Ca2+ influx through voltage‐dependent L‐type Ca2+ channels only in the beta‐cells in which glucose has already elevated [Ca2+]i, while high ACh increases [Ca2+]i at basal as well as stimulatory glucose concentrations by releasing Ca2+ from the ER. The former mechanism is likely to relate to the potentiator action and the latter to the initiator action of ACh on insulin release. High ACh and elevated glucose provoke both modes of Ca2+ signalling.
Role of nitric oxide in tachykinin‐induced increase in potential difference of rabbit tracheal mucosa.
Authors: Tamaoki J , Sakai A , Kondo M , Takemura H , Konno K
Published: December 2014

1. The effect of tachykinins on transepithelial potential difference (PD) of rabbit trachea and possible involvement of nitric oxide (NO) generation in vivo were investigated. 2. Perfusion of tracheal mucosa with neurokinin A (NKA) or substance P (SP) dose dependently increased PD in the presence of amiloride, with the potency being NKA > SP, but neurokinin B (NKB) had no effect. 3. Application of NG‐nitro‐L‐arginine methylester (L‐NAME, 10(‐3) M) attenuated the NKA‐induced increase in the amiloride‐sensitive PD, causing a rightward displacement of the dose‐response curve by approximately 1.0 log U, whereas NG‐nitro‐D‐arginine methylester (D‐NAME, 10(‐3) M) did not. 4. The inhibitory effect of L‐NAME was reversed by L‐arginine (10(‐2) M) but not by D‐arginine (10(‐2) M). 5. The release of NO was determined by a real‐time measurement of NO concentration ([NO]) in the perfusate using specific amperometric sensors for this molecule. 6. NKA and SP increased [NO] in a dose‐dependent manner, the maximal increase from the baseline value being 114 +/‐ 11 nM (mean +/‐ S.E.M., P < 0.001) and 54 +/‐ 6 nM (P < 0.01), respectively. 7. Histochemistry for NADPH diaphorase activity showed a strong staining within the epithelial cells. 8. We conclude firstly that tachykinins increase amiloride‐sensitive PD in vivo, which probably reflects Cl‐ movement from the submucosa toward the respiratory lumen in tracheal mucosa, and secondly that NO generation by epithelial cells may be involved in this process.
Detection of movements imposed on human hip, knee, ankle and toe joints.
Authors: Refshauge K M , Chan R , Taylor J L , McCloskey D I
Published: December 2014

1. The angular displacements necessary for 70% correct detection were determined in normal subjects at the hip, knee and ankle joints, and the interphalangeal joint of the big toe. Angular velocities between 0.1 and 50 deg s‐1 were tested. The hip and knee joints were tested in slight flexion and the ankle and toe joints were tested in the mid‐range of the normal excursion. The joints were carefully supported for testing and the muscles acting at the joints were relaxed. 2. When detection thresholds and velocities were assessed in terms of angular displacements and angular velocities, proprioceptive performances at the hip, knee and ankle joints were superior to that at the toe joint. 3. When detection levels and displacement velocities were expressed in terms of linear displacements and velocities at the tip of the extended toe for all four joints, instead of in angular terms, the ankle gave the best performance and the hip and knee the worst. 4. The detection level and velocity data were expressed also in terms of proportional changes in the fascicle lengths of muscles operating these joints. Analysis in these terms showed that performance was similar at the hip, knee and ankle joints, but that performance for the toe was much poorer than for the other joints. 5. These results for the hip, knee and ankle are similar to those previously measured for the elbow and distal interphlangeal joint of the finger, and are consistent with the theory that muscle fascicle length is the variable of significance to the central nervous system. However, the proprioceptive performance at the big toe is notably poorer than all other joints studied and analysed in terms of this variable.
Pre‐ and postsynaptic glutamate receptors at a giant excitatory synapse in rat auditory brainstem slices.
Authors: Barnes‐Davies M , Forsythe I D
Published: December 2014

1. Whole‐cell patch recordings were used to examine the EPSC generated by the calyx of Held in neurones of the medial nucleus of the trapezoid body (MNTB). Each neurone receives a somatic input from a single calyx (giant synapse). 2. A slow NMDA receptor‐mediated EPSC peaked in 10 ms and decayed as a double exponential with time constants of 44 and 147 ms. A fast EPSC had a mean rise time of 356 microseconds (at 25 degrees C), while the decay was described by a double exponential with time constants of 0.70 and 3.43 ms. 3. Cyclothiazide slowed the decay of the fast EPSC, indicating that it is mediated by AMPA receptors. The slower time constant was slowed to a greater extent than the faster time constant. Cyclothiazide potentiated EPSC amplitude, partly by a presynaptic mechanism. 4. The metabotropic glutamate receptor (mGluR) agonists, 1S,3S‐ACPD, 1S,3R‐ACPD and L‐2‐amino‐4‐phosphonobutyrate (L‐AP4) reversibly depressed EPSC amplitude. A dose‐response curve for 1S,3S‐ACPD gave an EC50 of 7 microM and a Hill coefficient of 1.2. 5. Analysis of the coefficient of variation ratio showed that the above mGluR agonists acted presynaptically to reduce the probability of transmitter release. Adenosine and baclofen also depressed transmission by a presynaptic mechanism. 6. alpha‐Methyl‐4‐carboxyphenylglycine (MCPG; 0.5‐1 mM) did not antagonize the effects of 1S,3S‐ACPD, while high concentrations of L‐2‐amino‐3‐phosphonopropionic acid (L‐AP3; 1 mM) and 4‐carboxy‐3‐hydroxyphenyglycine (4C3HPG; 500 microM) depressed transmission. 7. There was a power relationship between [Ca2+]o and EPSC amplitude with co‐operativity values ranging from 1.5 to 3.4. 8. The mechanism by which mGluRs modulate transmitter release appeared to be independent of presynaptic Ca2+ or K+ currents, since ACPD caused no change in the level of paired‐pulse facilitation or the duration of the presynaptic action potential (observed by direct recording from the terminal), indicating that the presynaptic mGluR transduction mechanism may be coupled to part of the exocytotic machinery. 9. Our data are not consistent with the presence at the calyx of Held of any one known mGluR subtype. Comparison of the time course and pharmacology of the fast EPSC with data from cloned AMPA receptors is consistent with the idea that GluR‐Do subunits dominate the postsynaptic channels.
Perinatal development of action potential propagation in cat rubrospinal axons.
Authors: Song W J , Okawa K , Kanda M , Murakami F
Published: December 2014

1. The development of action potential conduction was studied by intracellular recording of antidromic spikes in cat rubrospinal cells. 2. The distance between the C1 and L1 spinal segments increased linearly from 5.6 cm at embryonic day (E) 59 to 9.8 cm at postnatal day (P) 30. 3. The conduction time from the C1 segment to the rubrospinal neuron soma, estimated from antidromic spike latency evoked by stimulation of the C1 segment, decreased rapidly prior to birth and then slowly thereafter. This coincided with a reduction in conduction time variation between cells. 4. The conduction time from the red nucleus to the L1 segment followed a similar time course during development. The conduction time reached the adult value by P30, at which time the spinal cord is only half the adult length. 5. The conduction velocity between the C1 and L1 segments increased monotonically between E59 and P30, from a low of 1 m s‐1 to a maximum of 34 m s‐1. 6. The rise time of rubrospinal neuron somadendritic spikes followed a developmental time course similar to that for conduction time. 7. Myelination of rubrospinal axons, as judged by the presence of myelinated segment spikes, began to occur prior to E59. 8. These findings suggest that development of action potential propagation in rubrospinal cells can be divided into an early and a late stage: conduction time reaches the adult value during the early stage, i.e. by the first postnatal month, and is maintained during the late stage. We propose that myelination, axon diameter increase and maturation of membrane properties act to reduce conduction time to adult values during the early stage, while a proportional increase in fibre diameter with axonal length results in a constant conduction time during the late stage.
Atrial natriuretic factor release during pregnancy in rats.
Authors: Zhang Y , Novak K , Kaufman S
Published: December 2014

1. We investigated the control of atrial natriuretic factor (ANF) secretion during pregnancy. 2. Plasma ANF levels were measured in conscious virgin female rats under basal conditions, and after atrial distension with an indwelling balloon catheter. The rats were then mated, and the measurements repeated at 7, 14 and 21 days of pregnancy, and at 1 week postpartum. Plasma ANF levels were also measured in ovariectomized rats injected with progesterone, oestradiol, or oestradiol plus progesterone. 3. Basal plasma ANF levels were elevated at 7 and 14 days of pregnancy, but returned to prepregnant levels by 21 days. At 1 week postpartum, they were again elevated. 4. In response to atrial stretch, plasma ANF increased significantly in virgin rats (from 100 +/‐ 10 to 148 +/‐ 13 pg ml‐1, P < 0.001, n = 20). In contrast, there was no such secretory response observed in the pregnant and postpartum animals i.e. stretch‐induced secretion of ANF was markedly attenuated. 5. Treatment with exogenous oestradiol caused a significant increase in plasma ANF levels in acyclic rats. However, neither progesterone nor a combination of oestradiol plus progesterone had any effect. 6. It is concluded that basal and stretch‐induced ANF secretion are differentially influenced by pregnancy; oestradiol is identified as a potential stimulatory factor.
Characterization and regulation of a chloride channel from bovine tracheal epithelium.
Authors: Duszyk M , Liu D , Kamosinska B , French A S , Man S F
Published: December 2014

1. The patch‐clamp technique was used to characterize chloride channels from the apical membranes of bovine tracheal epithelial cells. Application of GTP gamma S or NaF to excised patches revealed the existence of a novel type of Cl‐ channel regulated by G‐proteins in a membrane‐delimited manner. 2. The channel had a linear current‐voltage relationship, with a conductance of 100‐120 pS. Its open probability was independent of voltage. 3. The channel was highly anion selective (permeability ratio, PNa/PCl = 0.06 +/‐ 0.04) and had the halide permeability sequence: I‐ > Br‐ > or = Cl‐ > F‐, corresponding to the Eisenman I sequence. This suggested that neither ionic size nor diffusion rate determined ion permeation through the channel. 4. The mole fraction behaviour was studied using fluoride and chloride ions. Mixtures of ions produced currents that would be expected from the linear combination of the two ions acting independently, indicating relatively simple permeation through the pore and compatible with a single ion binding site. 5. The channel was inhibited by the stilbene disulphonates SITS (4‐acetamido‐4'‐isothiocyanatostilbene‐2, 2'‐disulphonic acid) and DNDS (4,4'‐dinitrostilbene‐2,2'‐sulphonic acid). SITS introduced voltage dependence to channel gating and indicated the possible involvement of lysine residues in the channel permeation pathway. 6. NaF was unable to activate Cl‐ channels in the presence of the aluminum chelator, deferoxamine mesylate. This indicates that Al3+ ions play an important role in chloride channel activation by fluoride. NaF activation was not dependent on the presence of calcium ions. 7. The channel was insensitive to alkaline phosphatase and to the specific inhibitors of protein phosphatase types I and 2A, okadaic acid and calyculin A. 8. The channels could be activated by GTP gamma S or by NaF in the presence of the phospholipase A2 inhibitor quinacrine, indicating that this enzyme is not involved in channel regulation.