bluuebunny/crossref_metadata_2025 huggingface dataset:
Electrical resonance and Ca2+ influx in the synaptic terminal of depolarizing bipolar cells from the Goldfish retina
Authors: Burrone Juan , Lagnado Leon
Published: September 2004

Whole‐cell recordings and fura‐2 measurements of cytoplasmic [Ca2+] were made in depolarizing bipolar cells isolated from the retinae of goldfish. The aim was to study the voltage signal that regulates Ca2+ influx in the synaptic terminal.

The current‐voltage relation was linear up to about −44 mV. At this threshold, the injection of 1 pA of current triggered a maintained ‘all‐or‐none’ depolarization to a plateau of −34 mV, associated with a decrease in input resistance and a damped voltage oscillation with a frequency of 50–70 Hz and initial amplitude of 4–10 mV. A second frequency component of 5–10 Hz was often observed. In a minority of cells the response to current injection was transient, recovering with an undershoot.

Unstimulated bipolar cells generated similar voltage signals, driven by current entering the cell through a non‐specific cation conductance that continuously varied in amplitude.

The threshold for activation of the Ca2+ current was −43 mV and free [Ca2+]i in the synaptic terminal rose during a depolarizing response. Simultaneous measurements of the fluorescence associated with the membrane marker FM1–43 demonstrated that these Ca2+ signals stimulated exocytosis. Regenerative depolarizations and associated rises in [Ca2+]i were blocked by inhibiting l‐type Ca2+ channels with 30 μm nifedipine.

Depolarization beyond −40 mV also elicited an outwardly rectifying K+ current. Blocking this current by replacing external Ca2+ with Ba2+ caused the voltage reached during a depolarizing response to increase to +10 mV.

The majority of the K+ current was blocked by 100 nm charybdotoxin, indicating that it was carried by large‐conductance Ca2+‐activated K+ channels. A transient voltage‐gated K2+current remained, which began to activate at −40 mV. High‐frequency voltage oscillations were blocked by 100 nm charybdotoxin, but low‐frequency oscillations remained.

These results indicate that the voltage response of depolarizing bipolar cells is shaped by l‐type Ca2+ channels, Ca+‐activated K+ channels and voltage‐dependent K+ channels. This combination of conductances regulates Ca2+ influx into the synaptic terminal and confers an electrical resonance on the bipolar cell.
Dual actions of tetracaine on intramembrane charge in amphibian striated muscle
Authors: Huang Christopher L.H.
Published: September 2004

The effects of graded concentrations of tetracaine on the steady‐state and kinetic properties of intramembrane charge were examined in intact voltage‐clamped amphibian muscle fibres.

The micromolar tetracaine concentrations that were hitherto reported to abolish Ca2+ transients in skeletal muscle failed to affect significantly the steady‐state charge. Maximal reductions of such intramembrane charge required relatively high, 1–2 mm, concentrations of tetracaine.

The plots of maximum charge against tetracaine concentration suggested a saturable 1:1 drug binding that spared a fixed amount of tetracaine‐resistant (qβ) charge but inhibited a discrete fraction of susceptible (qγ) charge with a KD between 0.1 and 0.2 mm.

The qβ charge thus isolated by 2 mm tetracaine was conserved through a wide range of applied test voltages and pulse durations and regardless of whether the imposed transition from the holding potential (−90 mV) to the test potential took place in one or more steps.

Similarly, ‘on’ and ‘off’qβ currents that were elicited by voltage steps from fixed conditioning to varying test levels mapped onto non‐linear phase‐plane trajectories that nevertheless depended uniquely upon voltage. In contrast, the currents that followed voltage steps made from varying prepulse levels to fixed −90 or −20 mV test potentials mapped onto identical qβ phase‐plane trajectories that were independent of the prepulse history.

The charge movements that followed strong depolarizing voltage clamp steps to test potentials in the range −50 to 0 mV approximated simple monotonic decays that could empirically be described by a single time constant. Nevertheless, a complete inhibition of a tetracaine‐sensitive (qγ) charge movement by 2 mm tetracaine that left only qγ charge, sharply altered both the magnitude and the voltage dependence of these time constants. This establishes a distinct contribution of the qγ species to overall charge kinetics even at such test voltages.

Under such a criterion for the voltage dependence of charging kinetics, even the micromolar (0.05‐0.2mm) tetracaine concentrations that failed to markedly alter the steady‐state charge consistently increased the charging time constants yet did not influence their voltage sensitivity.

These findings demonstrate the existence of separate kinetic and steady‐state effects of tetracaine on intramembrane charge movements, at micromolar and millimolar anaesthetic concentrations, respectively. These parallel earlier effects of tetracaine that have been reported upon the transient and sustained components of sarcoplasmic reticular Ca2+ release. They also establish that maximally effective concentrations of tetracaine isolate a single distinct species of conserved (qβ) intramembrane charge.
A novel slow hyperpolarization‐activated potassium current (IK(SHA)) from a mouse hippocampal cell line
Authors: Wischmeyer Erhard , Karschin Andreas
Published: September 2004

A slow hyperpolarization‐activated inwardly rectifying K+ current (IK(SHA)) with novel characteristics was identified from the mouse embryonic hippocampus x neuroblastoma cell line HN9.10e.

The non‐inactivating current activated negative to a membrane potential of −80 mV with slow and complex activation kinetics (τact≈ 1–7 s) and a characteristic delay of 1–10 s (−80 to −140 mV) that was linearly dependent on the membrane potential.

Tail currents and instantaneous open channel currents determined through fast voltage ramps reversed at the K+ equilibrium potential (EK) indicating that primarily K+, but not Na+, permeated the channels.

I
K(SHA) was unaffected by altering the intracellular Ca2+ concentration between ∼0 and 10 μm, but was susceptible to block by 5 mm extracellular Ca2+, Ba2+ (K1= 0.42 mm), and Cs+ (K1= 2.77 mm)

In cells stably transformed with M2 muscarinic receptors, IK(SHA) was rapidly, but reversibly, suppressed by application of micromolar concentrations of muscarine.

At the single channel level K(SHA) channel openings were observed with the characteristic delay upon membrane hyperpolarization. Analysis of unitary currents revealed an inwardly rectifying I–V profile and a channel slope conductance of 7 pS. Channel activity persisted in the inside‐out configuration for many minutes.

It is concluded that IK(SHA) m HN9.10e cells represents a novel K+ current, which is activated upon membrane hyperpolarization. It is functionally different from both classic inwardly rectifying Ikir currents and other cationic hyperpolarization‐activated 7H currents that have been previously described in neuronal or glial cells.
Synapse Formation and Hypoxic Signalling in Co‐Cultures of Rat Petrosal Neurones and Carotid Body Type 1 Cells
Authors: Zhong Huijun , Zhang Min , Nurse Colin A.
Published: September 2004

To investigate synaptic mechanisms mediating chemosensory signalling in the carotid body, we developed co‐cultures of chemoreceptor type 1 cell clusters and dissociated petrosal neurones (PNs) from 7‐ to 14‐day‐old rat pups and tested for functional connectivity in CO2–HCO3− ‐or Hepes‐buffered medium at ∼35 °C.

When cultured without type 1 cells, PNs were almost always quiescent (n= 104) and unresponsive to hypoxia (PO2= 5–25 mmHg) during perforated patch, whole‐cell recordings of membrane potential or voltage‐activated currents; in contrast, many PNs (77 out of 170) that were juxtaposed to type 1 cell clusters in co‐culture displayed spontaneous activity, comprising spikes and subthreshold potentials (SSPs) that resembled synaptic potentials.

Additional tests suggested that de novo chemical synapses developed between PNs and type 1 cell clusters in vitro. For example: (i) the spontaneous activity was reversibly suppressed by substituting low calcium‐high magnesium in the bath; (ii) SSPs had variable amplitudes and persisted following action potential blockade with TTX (1 μm); (iii) the interval distribution between successive spontaneous events appeared random; and (iv) the frequency of spontaneous potentials was diminished (reversibly) by the nicotinic antagonist hexamethonium (100 /tM), suggesting contributions from the spontaneous release of ACh.

Many complexes of ‘juxtaposed’ PNs and type 1 clusters were physiologically functional, since exposure to hypoxia caused a reversible depolarization and/or increased spike discharge in ∼30% of such neurones (n= 140). The hypoxia‐induced spike discharge persisted in the presence of the dopamine D2 receptor blocker spiperone (10–50 μm; n= 5); however, this discharge was reversibly inhibited by 100–200 μm hexamethonium, suggesting that it was mediated, at least in part, by ACh acting through nicotinic receptors.

The hypoxia‐induced spike discharge and frequency of spontaneous potentials in co‐cultured PNs were reversibly suppressed when the buffer was switched from C02‐HC03∼ to Hepes (10 mm) at pH 7.4;′further, ‘functional’ PNs that displayed spontaneous activity and/or hypoxia‐induced responses in co‐culture were encountered more frequently in CO2–HCO3− (≥ 40 %) than in Hepes (≤26 %) buffer.

We conclude that functional chemical synapses can develop de novo in cultures of carotid body type 1 cells and PNs and that ACh is probably an important excitatory neuro‐transmitter secreted from type 1 cells during hypoxic chemotransduction in the rat carotid body.
Functional nicotinic ACh receptors on interneurones in the rat hippocampus
Authors: Jones Susan , Yakel Jerrel L.
Published: September 2004

Neuronal nicotinic ACh receptors (nAChRs) were studied in the rat hippocampal slice preparation using whole‐cell patch‐clamp recording techniques.

Responses to ACh (100 μm) were detected on inhibitory interneurones in the CA1 field of the hippocampus proper and in the dentate gyrus, but not on principal excitatory neurones in either region. The different neuronal types were identified based on their morphology and location.

ACh excited interneurones in the hippocampus and dentate gyrus in current‐clamp recordings. In voltage‐clamp recordings, ACh‐activated inward currents were recorded from interneurones in the presence of blockers of synaptic transmission and the muscarinic ACh receptor antagonist atropine. The zero current potential for this response to ACh was near 0 mV.

The effect of ACh was mimicked by the nAChR‐selective agonists nicotine (100 μm) and 1,1‐dimethyl‐4‐phenyl‐piperazinium iodide (DMPP, 100 μm). The response to ACh was reversibly antagonized by the neuronal nAChR antagonist mecamylamine (10 μm). The nAChR α7 subunit‐selective antagonists α‐bungarotoxin (100 nm) and methyllycaconitine (10 nm) also inhibited the response to ACh.

These observations demonstrate the presence of functional nAChRs on inhibitory interneurones in the rat hippocampus. Thus, a novel mechanism by which ACh can regulate neuronal activity in the hippocampus is revealed.
Calcium action potentials restricted to distal apical dendrites of rat neocortical pyramidal neurons
Authors: Schiller Jackie , Schiller Yitzhak , Stuart Greg , Sakmann Bert
Published: September 2004

Simultaneous whole‐cell voltage and Ca2+ fluorescence measurements were made from the distal apical dendrites and the soma of thick tufted pyramidal neurons in layer 5 of 4‐week‐old (P28–32) rat neocortex slices to investigate whether activation of distal synaptic inputs can initiate regenerative responses in dendrites.

Dual whole‐cell voltage recordings from the distal apical trunk and primary tuft branches (540–940 μm distal to the soma) showed that distal synaptic stimulation (upper layer 2) evoking a subthreshold depolarization at the soma could initiate regenerative potentials in distal branches of the apical tuft which were either graded or all‐or‐none. These regenerative potentials did not propagate actively to the soma and axon.

Calcium fluorescence measurements along the apical dendrites indicated that the regenerative potentials were associated with a transient increase in the concentration of intracellular free calcium ([Ca2+]i) restricted to distal dendrites.

Cadmium added to the bath solution blocked both the all‐or‐none dendritic regenerative potentials and local dendritic [Ca2+]i transients evoked by distal dendritic current injection. Thus, the regenerative potentials in distal dendrites represent local Ca2+ action potentials.

Initiation of distal Ca2+ action potentials by a synaptic stimulus required coactivation of AMPA‐ and NMDA‐type glutamate receptor channels.

It is concluded that in neocortical layer 5 pyramidal neurons of P28–32 animals glutamatergic synaptic inputs to the distal apical dendrites can be amplified via local Ca2+ action potentials which do not reach threshold for axonal AP initiation. As amplification of distal excitatory synaptic input is associated with a localized increase in [Ca2+]i these Ca2+ action potentials could control the synaptic efficacy of the distal cortico‐cortical inputs to layer 5 pyramidal neurons.
Sarcomere length dependence of the rate of tension redevelopment and submaximal tension in rat and rabbit skinned skeletal muscle fibres
Authors: McDonald Kerry S. , Wolff Matthew R. , Moss Richard L.
Published: September 2004

We examined the hypothesis that in skeletal muscle the steep relationship between twitch tension and sarcomere length (SL) within the range 2.30 to 1.85 μm involves SL‐dependent alterations in the rate of tension development.

In skinned preparations of both rat slow‐twitch and rabbit fast‐twitch skeletal muscle fibres the rate of tension redevelopment (ktr) at 15°C was reduced at short SL (∼2.00/μm) compared with a longer SL (∼2.30 μm). In submaximally activated fibres, the decrease in ktr over this range of lengths was greater in fast‐twitch fibres (38% reduction) than in slow‐twitch fibres (14% reduction).

Ca2+ sensitivity of tension, as assessed as the pCa (−log[Ca2+]) for half‐maximal activation, or pCa50, decreased to a greater extent in rabbit fast‐twitch skeletal muscle fibres than in slow‐twitch fibres from both rabbit and rat when SL was reduced from ∼2.30 to ∼1.85 μm. The ΔpCa50 over this SL range was 0.24 ± 0.07 pCa units in fast‐twitch fibres from rabbit psoas muscle. The ΔpCa50 for slow‐twitch fibres from rabbit and rat soleus muscle was 0.08 ± 0.02 and 0.10 ± 0.04 pCa units, respectively.

Osmotic compression of both slow‐twitch and fast‐twitch fibres at a SL of 2.00 μm increased ktr to values similar to those obtained at a SL of 2.30 μm in the absence of dextran. This result indicates that the slower rate of tension redevelopment at short SL is due in large part to the increase in interfilament lattice spacing associated with shorter SL.

Taken together, these results suggest that length dependence of twitch tension is, in part, due to length dependence of isometric cross‐bridge interaction kinetics, an effect that is mediated by length‐dependent changes in interfilament lattice spacing.
Rhythmic bursting of pre‐ and post‐inspiratory neurones during central apnoea in mature mice
Authors: Paton Julian F. R.
Published: September 2004

Stimulation of pulmonary vagal C fibres (PCFs) inhibits inspiration but the response pattern of respiratory rhythm‐generating neurones is unknown. This study provides the first description of the effects of PCF stimulation on six different types of respiratory neurones located in the ventrolateral medulla of the mature mouse.

Studies were performed in both urethane‐anaesthetized (1.5 g kg−1 I.P.) mature mice and in an arterially perfused working heart‐brainstem preparation (WHBP). In both preparations the respiratory motor pattern of phrenic and recurrent laryngeal nerves were comparable. Stimulation of PCFs, using phenylbiguanide (2–5 μg) injected into the right atrium, evoked a similar respiratory and cardiac response pattern in both anaesthetized and perfused mice, which included: (i) a significant prolongation of the inter‐inspiratory interval; (ii) an increase in the duration and amplitude of post‐inspiratory (ii) activity; and (iii) an atropine‐sensitive bradycardia (50–260 beats min−1).

In the WHBP, PCF stimulation evoked a depolarization (11 ± 1 mV) and high frequency tonic discharge (up to 64 Hz) in ten out of twenty‐one PI neurones. During the PCF‐induced prolongation of PI activity all other PI neurones (n= 11), as well as pre‐inspiratory neurones (PreI; n= 11), displayed oscillations in membrane potential and/or rhythmic bursting at a similar frequency of 0.7‐1.0 Hz. Other respiratory neurones recorded, including stage II expiratory neurones (n= 7), early‐ (n= 6), ramp‐ (n= 16) and late‐inspiratory neurones (n= 4), ceased firing rhythmically during PCF stimulation.

The firing behaviour of PI and PreI neurones was assessed after switching to a low Ca2+ (0.2 mm)‐high Mg2+ (5.25 mm) perfusate to block synaptic transmission in the WHBP. In the absence of synaptic transmission, PreI neurones (n= 7/8) continued to discharge rhythmically, whereas all other respiratory cell types (including PI neurones, n= 5) fired tonically.

In conclusion, stimulation of PCFs elicits a reflex‐evoked prolongation of the PI phase of the respiratory cycle and excitation of PI neurones including rhythmic discharging. It is suggested that this rhythmic bursting depends on inhibitory connections from PreI neurones. The functional significance of these central ‘apnoeic rhythms’ are discussed.
Rhythmic motor activity and interlimb co‐ordination in the developing pouch young of a wallaby (Macropus eugenii)
Authors: Ho Stephen M.
Published: September 2004

The forelimb motor behaviour of developing wallaby was studied. A clock‐like alternating movement was reactivated whenever the animal was removed from the pouch.

Forelimb stepping frequency increased during the first 3 weeks of development, while the phase relationship remained constant. Forelimb activity could be affected by altering the afferent feedback from the contralateral limb, or an increase in ambient temperature.

In vitro experiments were performed using an isolated brainstem‐spinal cord preparation from animals up to 6 weeks postnatal. Fictive locomotor activity could be evoked by electrical stimulation or bath‐applied NMDA (< 10 μm).

Bath‐applied strychnine (10‐25 μm) and bicuculline (10‐50 μm) disrupted the phase relationship between motor pools, while rhythmic motor discharge remained in the absence of these inhibitory pathways.

The present findings indicate that the pattern generator that underlies the robust forelimb movement during the first journey to the pouch is retained for different motor functions during in‐pouch development. The neural network that underlies such behaviour can be divided into two major components, a rhythm generator within each hemicord, and a pattern co‐ordinating pathway which involve both glycinergic and GABAergic interneurones.
Novel glutamate‐ and GABA‐independent synaptic depolarization in granule cells of guinea‐pig hippocampus
Authors: Forti Matteo , Michelson Hillary B.
Published: September 2004

Dual intracellular recordings of granule cells, hilar interneurons and CA3 pyramidal cells were performed in transverse slices of guinea‐pig hippocampus. At resting membrane potential, in the presence of 4‐aminopyridine, ionotropic glutamate receptor antagonists and the GABAA receptor antagonist bicuculline, granule cells showed spontaneous, large amplitude depolarizations correlated with synchronous bursting activity of interneurons.

Under these conditions, pyramidal cells exhibited large amplitude monophasic GABAB inhibitory postsynaptic potentials (IPSPs) synchronous with the GABAergic interneuron burst discharges. The granule cells also received a GABAB input, which was evident only when the neurons were depolarized by DC injection. The GABAB receptor antagonist CGP 55845A (CGP) blocked the GABAB IPSPs in both pyramidal cells and granule cells; however, the depolarizing potential in granule cells was unaffected by the drug.

The granule cell depolarization in the presence of CGP was monophasic and exhibited linear voltage dependence with a reversal potential around −40 mV, suggesting that it was generated by a synaptic input activating a mixed cationic current.

The granule cell depolarization was abolished following the addition of tetrodotoxin to the bath. In addition, perfusing the slice with a low Ca2+‐containing solution (0.5 mm Ca2+–10 mm Mg2+) also abolished the granule cell depolarization, confirming the synaptic origin of the event.

(S)‐Methyl‐4‐carboxyphenylglycine, l‐(+)‐2‐amino‐3‐phosphonopropionic acid, propranolol and atropine did not affect the granule cell depolarization, indicating that metabotropic glutamate receptors,β‐adrenergic receptors and muscarinic cholinergic receptors were not involved in generating the granule cell depolarizing synaptic response.

These findings indicate that, in the absence of both glutamatergic and GABAergic inputs, synchronous interneuronal activity can produce a depolarizing synaptic response in granule cells. The neurochemical responsible for the depolarization is currently under investigation.
Characterization of the inward current induced by metabotropic glutamate receptor stimulation in rat ventromedial hypothalamic neurones
Authors: Lee K. , Boden P. R.
Published: September 2004

Whole‐cell patch clamp recordings were made from rat ventromedial hypothalamic neurones in slices of brain tissue in vitro. Bath application of 50 μm (1S, 3R)‐1‐aminocyclopentane‐1,3‐dicarboxylic acid (1S,3R‐ACPD) depolarized all neurones tested by activation of an inward current of approximately 55 pA at −60 mV.

The inward current elicited by 1S,3R‐ACPD was unaffected by K+ channel blockade with external Cs+, Ba2+ or TEA. However, the current was significantly reduced by replacement of the external NaCl with either Tris‐HCl or LiCl.

The 1S,3R‐ACPD‐induced current was reduced by the heavy metal ions Ni2+ or La3+ and also by the Na+–Ca2+ exchange current inhibitor 3′,4′‐dichlorobenzamil.

The effects of 1S,3R‐ACPD were mimicked by the group I metabotropic agonist 3,5‐dihydroxyphenylglycine (DHPG) but not by the group III selective agonist, l‐2‐amino‐4‐phosphonobutanoate (l‐AP4). Furthermore, the effects of 1S,3R‐ACPD were inhibited by the metabotropic antagonists α‐methyl‐4‐carboxyphenylglycine (MCPG) and 1‐aminoindan‐1,5‐dicarboxylic acid (AIDA) but not by the presynaptic metabotropic receptor antagonists α‐methyl‐4‐phosphonophenylglycine (MPPG) or α‐methyl‐4‐tetrazolylphenylglycine (MTPG).

Photorelease of caged GDPβS inside neurones irreversibly blocked the 1S,3R‐ACPD‐induced current whilst photolysis of caged GTPγS inside neurones irreversibly potentiated this current.

The PLC inhibitor U‐73122 significantly reduced the size of the inward current induced by 1S,3R‐ACPD. This effect was not mimicked by the inactive analogue U‐73343.

Flash photolysis of the caged calcium chelator diazo‐2 inside neurones diminished the response to 1S,3R‐ACPD.

It is concluded that group I metabotropic glutamate receptors depolarize neurones in the VMH by activation of a Na+–Ca2+ exchange current through a G‐protein coupled increase in intracellular Ca2+.
Distribution of sarcomere length and intracellular calcium in mouse skeletal muscle following stretch‐induced injury
Authors: Balnave C. D. , Davey D. F. , Allen D. G.
Published: September 2004

The effect on sarcomere organization of stretching intact single skeletal muscle fibres by 50% of their optimum length (Lo) during ten consecutive short tetani was investigated. Stretch reduced tetanic force to 36 ± 4% of the pre‐stretch condition. Sarcomere organization was analysed using both electron and confocal microscopy. For confocal microscopy the striation pattern was examined by fluorescently staining F‐actin with rhodamine–phalloidin.

Electron microscopy revealed that fibres which had been stretched during contraction contained areas of severe sarcomere disorganization, as well as adjacent sarcomeres of normal appearance.

Confocal images of stretched fibres, which had been fixed and stained with rhodamine–phalloidin, showed focal regions of overstretched sarcomeres and regions where sarcomeres of adjacent myofibrils were out of alignment with each other. Analysis of all sarcomeres along the length of fibres showed regions of sarcomere inhomogeneity were distributed throughout the fibre length and cross‐section.

Fibres were microinjected with the fluorescent [Ca2+]i indicator fura‐2 before being stretched. Conventional wide‐field fluorescence imaging microscopy showed that the tetanic [Ca2+]i was reduced after stretching but remained uniformly distributed.

This study confirms the finding that stretch‐induced muscle injury has components caused by disorganization of the myofibrillar array and by failure of tetanic Ca2+ release. The structural damage is spatially heterogeneous whereas the changes in Ca2+ release appear to be spatially homogeneous.
Changes in initiation of orienting gaze shifts after muscimol inactivation of the caudal fastigial nucleus in the cat
Authors: Goffart Laurent , Pélisson Denis
Published: September 2004

The production of a goal‐directed saccadic gaze shift involves the specification of movement amplitude and direction, and the decision to trigger the movement. Behavioural and neurophysiological data suggest that these two functions involve separate processes which may interact.

The medio‐posterior cerebellar areas are classically assigned a major contribution to the control of saccade metrics, and previous cerebellar lesion studies have revealed marked dysmetria of visually triggered gaze shifts. In contrast, these studies did not provide evidence for a cerebellar role in saccadic initiation.

In the present study, we investigated in the head‐unrestrained cat the deficits in both the initiation and the metrics control of saccadic gaze shifts following pharmacological inactivation of the caudal part of the fastigial nucleus (cFN).

After cFN inactivation, latencies for contraversive gaze shifts increased to about 137 ± 28 % of normal, and latencies for ipsiversive gaze shifts decreased to about 84 ± 8 % of normal. Similar changes in head movement latency were observed, such that the temporal coupling between eye and head components remained largely unaffected.

Contraversive gaze shifts were more hypometric as their latency increased. In contrast, the degree of hypermetria in ipsiversive gaze shifts was unrelated to latency.

These results suggest a functional role of the medio‐posterior cerebellum in gaze shift initiation and in storing information about the target location and/or the desired gaze shift amplitude.
Differing effects of histamine and serotonin on microvascular permeability in anaesthetized rats
Authors: Michel C. C. , Kendall S.
Published: September 2004

We have investigated simultaneous changes in the hydraulic permeability (Lp) and the retention of perfusate macromolecules in single mesenteric venules of anaesthetized rats during perfusion with either histamine or serotonin.

The venules were microperfused in situ. Retention of macromolecules was assessed from the effective oncotic pressure (σΔπ) exerted by the perfusate across the vessel walls. Lp and σΔπ were estimated by the red cell microperfusion technique.

Perfusion with histamine (at concentrations between 16 μm and 3.26 mm) and serotonin (at concentrations between 26 μm and 1.3 mm) transiently increased Lp and reduced σΔπ Maximal changes were seen at 6–9 min with histamine and at 3 min with serotonin.

Maximal increases in Lp were greater with histamine (approximately 3 ‐fold) than with serotonin (1.5‐ to 2‐fold). Serotonin, however, decreased σΔπ from a baseline of 14–15 cm H2O to one of 6–7 cm H2O whereas the fall of σΔπ with histamine was only from 14–15 cm H2O to 10–11 cm H2O.

The data are consistent with the hypothesis that serotonin increases permeability by inducing openings in the venular endothelium which do not retain macromolecules. If histamine also increases permeability by gap formation, these gaps are able to retain macromolecules to a significant extent.
Dynamics of viscoelastic properties of rat cardiac sarcomeres during the diastolic interval: involvement of Ca2+
Authors: Stuyvers Bruno D. M. Y. , Miura Masahito , Keurs Henk E. D. J. ter
Published: September 2004

Cardiac sarcomere stiffness was investigated during diastole in eighteen trabeculae dissected from the right ventricle of rat heart. The trabeculae were stimulated at 0.5 Hz, in a modified Krebs–Henseleit solution (pH, 7.4; 25 °C). Sarcomere length (SL) was measured using high resolution (±2 nm) laser diffraction techniques. Force (F) was measured with a silicon strain gauge.

SL increased exponentially (amplitude, 25 ± 9 nm; n= 5) throughout diastole. This increase occurred even at slack SL, showing that this phenomenon was due to an internal expansion. The majority of the muscles showed discrete spontaneous fluctuations of SL (amplitude < 20 nm) starting ∼l s after the end of the twitch.

The intracellular free Ca2+ concentration ([Ca2+]i) was measured from the fluorescence of microinjected fura‐2 salt in seven trabeculae under the same experimental conditions. [Ca2+]i continuously declined (from 240 to 90 nm) during diastole following a monoexponential time course (time constant, 210–325 ms).

The stiffness of the sarcomere was evaluated at 10, 30, 50, 70 and 90% of diastole using bursts (30 ms) of 500 Hz sinusoidal perturbations of muscle length (amplitude of SL oscillations < 30 nm). At 1 mm external Ca2+ concentration ([Ca2+]o), the average stiffness modulus (Mod) increased from 9.3 ± 0.6 to 12 ± 0.6 mN mm−2μm1 (n= 18; P < 0.05), while the average phase shift (Ф) between F and SL signals decreased from 84 ± 3 to 73 ± 4 deg (n= 18; P < 0.05) between 10 and 90% during diastole. The increase in Mod and the decrease in Ф reversed when spontaneous activity occurred. When [Ca2+]0 was raised to 2 mm, the stiffness time course reversed approximately 450 ms earlier, simultaneously with the occurrence of spontaneous activity.

Our results show that diastole is only an apparent steady state and suggest that the structural system responsible for the viscoelastic properties of the sarcomere is regulated by [Ca2+]i in the submicromolar range. Different possible origins of the dynamic changes in viscoelasticity during diastole are discussed.
Age‐related functional changes of the glutamate receptor channels in rat Meynert neurones
Authors: Akaike Norio , Rhee Jeong‐Seop
Published: September 2004

The developmental changes of glutamate receptors (GluRs) in acutely dissociated rat Meynert neurones were investigated using the conventional whole‐cell and nystatin perforated patch recording modes under voltage‐clamp conditions.

The neurones became less responsive to N‐methyl‐d‐aspartic acid (NMDA) with age, most dramatically between 1 day and 2 weeks, while the responses to kainic acid (KA) and l‐α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA) gradually increased. The metabotropic GluR response appeared a few days after birth, but thereafter no further change was observed.

The decrease in the NMDA response during postnatal development was due to an abrupt reduction in the number of receptors without affecting the affinity, voltage‐dependent Mg2+ blockade or high Ca2+ permeability (Pca/PCs≈ 7.0).

P
ca/PCs in the presence of KA decreased from 2.8 in the 1‐day‐old (1D) rat neurones to 1.1 and 0.44 in the 2‐week‐old (2W) and 6‐month‐old (6M) rat neurones, respectively. The concentration–response relationship for KA shifted to the left with age. The KA response was not affected by NS‐102, a KA‐selective antagonist, thus indicating that the increased affinity of the receptor for the ligand resulted from the change in the AMPA receptor channel subunits.

The AMPA response in the presence of 10−4 M cyclothiazide showed a change in the inward rectifying current–voltage relationship with age. The KA response was strongly cross‐desensitized by the addition of AMPA and was also blocked by 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX), whereas a rapid desensitization of the AMPA response was removed in a concentration‐dependent manner by cyclothiazide. These results indicate that the non‐NMDA receptor channels are assembled from the subunits of the AMPA receptor family without the GluR‐2 subunit, thus resulting in a high Ca2+ permeability.

The l‐glutamate (Glu)‐induced responses were more sensitive to dl‐2‐amino‐5‐phosphonopentanoic acid (APV) in the 1D rat neurones than in the adult rat neurones.

Both NMDA and KA raised the intracellular Ca2+ concentration ([Ca2+]i) in all neurones of 1D, 2W and 6M rats, though the charybdotoxin‐sensitive Ca2+‐activated K+ current (IK(ca)) did not appear in the 1D rat neurones. An age‐related prolongation of both IK(Ca) decay and [Ca2+]i clearance was also seen after the removal of KA.

It was thus concluded that the age‐related changes of ionotropic receptors appear to play a key role in the activities of immature and mature rat Meynert cholinergic neurones. The KA‐induced IK(Ca), which developed with ageing, may thus function as one of the negative feedback systems, and thereby prevent excess cell excitation and neural damage, especially in adult rats.
Ionic currents during sustained pacemaker activity in rabbit sino‐atrial myocytes
Authors: Zaza Antonio , Micheletti Maria , Brioschi Angelica , Rocchetti Marcella
Published: September 2004

The contribution of various ionic currents to diastolic depolarization (DD) in rabbit sino‐atrial myocytes was evaluated by the action potential clamp technique. Individual currents were identified, during sustained pacemaking activity reproduced under voltage clamp conditions, according to their sensitivity to supecific channel blockers.

The current sensitive to dihydropyridines (DHPs), blockers of L‐type Ca2+ current (ICa,L), was small and outward during most of DD. Diastolic DHP‐sensitive current was affected by changes in the driving force for K+, but it was insensitive to E‐4031, which blocks the current termed IK,r; it was abolished by cell dialysis with a Ca2+ chelator.

The current sensitive to 2 mm Cs+ (ICs), a blocker of hyperpolarization‐activated current (If), was inward during the whole DD and it was substantially larger than the net inward current flowing during this phase. However, diastolic IK,r, identified in the same cells as the current sensitive to the blocker E‐4031, exceeded ICs 2‐fold.

These findings suggest that: (a) Ca2+ influx during the pacemaker cycle increases a K+ conductance, thus inverting the direction of the net current generated by L‐type Ca2+ channel activity during DD; (b) the magnitude of If would be adequate to account fully for DD; however, the coexistence of a larger IK,r suggests that other channels besides If contribute inward current during this phase.
Anaplerotic processes in human skeletal muscle during brief dynamic exercise
Authors: Gibala Martin J. , MacLean Dave A. , Graham Terry E. , Saltin Bengt
Published: September 2004

This study examined changes in tricarboxylic acid cycle intermediates (TCAIs) in human skeletal muscle during 5min of dynamic knee extensor exercise (∼80% of maximum workload) and following 2 min of recovery.

The sum of the seven measured TCAIs (ΣTCAIs) increased from 1.10 ± 0.08mmol (kg dry weight)−1 at rest to 3.12 ± 0.24, 3.86 ± 0.35 and 4.33 ± 0.30 mmol (kg dry weight)−1after 1, 3 and 5 min of exercise, respectively (P≤ 0.05). The ΣTCAIs after 2 min of recovery (3.74 ± 0.43 mmol (kg dry weight)−1) was not different compared with 5 min of exercise.

The rapid increase in ΣTCAIs during exercise was primarily mediated by large changes in succinate, malate and fumarate. These three intermediates accounted for > 90 % of the net increase in ΣTCAIs during the first minute of contraction.

Intramuscular alanine increased after 1 min of exercise by an amount similar to the increase in the ΣTCAIs (2.33 mmol (kg dry weight)−1) (P≤ 0.05). Intramuscular pyruvate was also higher (P≤0.05) during exercise, while intramuscular glutamate decreased by ∼50% within 1 min and remained low despite an uptake from the circulation (P≤ 0.05).

The calculated net release plus estimated muscle accumulation of ammonia after 1 min of exercise (∼60 μmol (kg wet weight)−1) indicated that only a minor portion of the increase in ΣTCAIs could have been mediated through the purine nucleotide cycle and/or glutamate dehydrogenase reaction.

It is concluded that the close temporal relationship between the increase in ΣTCAIs and changes in glutamate, alanine and pyruvate metabolism suggests that the alanine amino‐transferase reaction is the most important anaplerotic process during the initial minutes of contraction in human skeletal muscle.
Enhanced pulmonary and active skeletal muscle gas exchange during intense exercise after sprint training in men
Authors: McKenna Michael J. , Heigenhauser George J. F. , McKelvie Robert S. , Obminski George , MacDougall J. Duncan , Jones Norman L.
Published: September 2004

This study investigated the effects of 7 weeks of sprint training on gas exchange across the lungs and active skeletal muscle during and following maximal cycling exercise in eight healthy males.

Pulmonary oxygen uptake (V̇O2) and carbon dioxide output (V̇CO2) were measured before and after training during incremental exercise (n= 8) and during and in recovery from a maximal 30 s sprint exercise bout by breath‐by‐breath analysis (n= 6). To determine gas exchange by the exercising leg muscles, brachial arterial and femoral venous blood O2 and CO2 contents and lactate concentration were measured at rest, during the final 10 s of exercise and during 10 min of recovery.

Training increased (P < 0.05) the maximal incremental exercise values of ventilation (V̇E, by 15.7 ± 7.1%), V̇CO2 (by 9.3 ± 2.1%) and V̇O2 (by 15.0 ± 4.2%). Sprint exercise peak power (3.9 ± 1.0% increase) and cumulative 30 s work (11.7 ± 2.8% increase) were increased and fatigue index was reduced (by −9.2 ± 1.5%) after training (P < 0.05). The highest V̇E, V̇CO2 and V̇O2 values attained during sprint exercise were not significantly changed after training, but a significant (P < 0.05) training effect indicated increased V̇E(by 19.2 ± 7.9%), V̇CO2(by 9.3 ± 2.1%) and V̇O2(by 12.7 ± 6.5%), primarily reflecting elevated post‐exercise values after training.

Arterial O2 and CO2 contents were lower after training, by respective mean differences of 3.4 and 21.9 ml l−1 (P < 0.05), whereas the arteriovenous O2 and CO2 content differences and the respiratory exchange ratio across the leg were unchanged by training.

Arterial whole blood lactate concentration and the net lactate release by exercising muscle were unchanged by training.

The greater peak pulmonary V̇O2 and V̇CO2 with sprint exercise, the increased maximal incremental values, unchanged arterial blood lactate concentration and greater sprint performance all point strongly towards enhanced gas exchange across the lungs and in active muscles after sprint training. Enhanced aerobic metabolism after sprint training may contribute to reduced fatigability during maximal exercise, whilst greater pulmonary CO2 output may improve acid‐base control after training.
Modulation of stimulus—secretion coupling in single rat gonadotrophs
Authors: Thomas Paul , Waring Dennis W.
Published: September 2004

Exocytosis and intracellular [Ca2+] were determined simultaneously in single anterior pituitary gonadotrophs from ovariectomized female rats. Dispersed cells were cultured for 2–4 days with or without 0.2 nm oestradiol‐17 β (E2) before use. Cells were stimulated with either gonadotrophin releasing hormone (GnRH) or by membrane depolarization. Exocytosis was determined from the change in membrane capacitance (Cm) using the perforated‐patch whole‐cell recording technique. Intracellular [Ca2+] was measured using fura‐2 fluorescence.

The exocytotic response to 1 nm GnRH was characterized by a wide spectrum of responses, ranging from exocytotic bursts to relatively slow, graded increases that were dependent on the evoked intracellular Ca2+ pattern. A kinetic model is presented that incorporates the observed steep dependence of exocytosis on measured intracellular [Ca2+]; simulated exocytosis reasonably approximated observed exocytotic responses, both kinetically and quantitatively. The model also suggests that the modulatory effects of E2 are brought about either by a change in the Ca2+ sensitivity of exocytosis or by a preferential clustering of docked‐secretory granules close to sites of Ca2+ release. The results suggest that in gonadotrophs an oscillatory Ca2+ signal is sensed by the exocytotic apparatus in a modified form of digital encoding.

Exocytosis in E2‐treated cells was 3‐fold greater than in non‐treated cells for GnRH‐evoked secretion, and 38% greater for depolarization; however, there was no effect of E2 on the intracellular Ca2+ response to either stimulus. The results show that maximum expression of the effect of E2 on exocytosis requires activation of GnRH‐dependent pathways.
Extracellular glucose turnover in the striatum of unanaesthetized rats measured by quantitative microdialysis
Authors: Fray Anne E. , Boutelle Martyn , Fillenz Marianne
Published: September 2004

Steady‐state and time‐resolved quantitative microdialysis was used to measure dialysate concentration, extracellular concentration and the in vivo recovery of glucose in rat striatum.

The extracellular concentration of glucose, determined by the zero net flux method of Lönnroth, was 350 ± 20 μm and the in vivo recovery was 39 ± 2%.

Veratridine caused a steep decrease in dialysate glucose after an initial delay of 7.5 min. When steady‐state glucose levels had been reached in the presence of veratridine the extracellular concentration was reduced to zero, but there was no significant change in in vivo recovery.

Measurement of the dynamic changes during the administration of veratridine showed an immediate decrease in extracellular glucose concentration and a steep rise in in vivo recovery, which accounted for the delay in the decrease in dialysate glucose. When extracellular concentration reached zero, in vivo recovery returned to control levels.
On the nature of anomalous rectification in thalamocortical neurones of the cat ventrobasal thalamus in vitro
Authors: Williams Stephen R. , Turner Jonathan P. , Hughes Stuart W. , Crunelli Vincenzo
Published: September 2004

Intracellular sharp electrode current clamp and discontinuous single electrode voltage clamp recordings were made from thalamocortical neurones (n= 57) of the cat ventrobasal thalamus in order to investigate the mechanisms underlying anomalous rectification.

Under current clamp conditions, voltage‐current (V–I) relationships in a potential range of −55 to −110 mV demonstrated anomalous rectification with two components: fast rectification, which controlled the peak of negative voltage deviations, and time‐dependent rectification. Time‐dependent rectification was apparent as a depolarizing sag generated during the course of negative voltage deviations, was first formed at potentials in the range −60 to −70 mV, and was sensitive to 3 mm Cs+ (n= 6). Similarly, under voltage clamp conditions, instantaneous and steady‐state I–V relationships demonstrated anomalous rectification. A slowly activating inward current with an activation threshold in the range of −65 to −70 mV formed time‐dependent rectification. This current was sensitive to Cs+ (3 mm) (n= 3) and had properties similar to the slow inward mixed cationic current (Ih).

4‐(N‐Ethyl‐N‐phenylamino)‐l,2‐dimethyl‐6‐(methylamino)‐pyrimidinium chloride (ZD7288) (100–300 μm) irreversibly blocked time‐dependent rectification mediated by Ih (n= 23 of 25 neurones), and led to a hyperpolarization of the resting membrane potential (6.8 ± 0.5 mV). In the presence of ZD 7288, V–I and I–V relationships exhibited fast anomalous rectification, first activated from potential more negative than −80 mV.

Ba2+ (100 μm) (n= 8), in the continuous presence of ZD 7288, reversibly linearized peak V–I and instantaneous I–V relationships over a potential range of −70 to −120 mV, and led to a membrane depolarization (13.3 ± 4.2 mV) or tonic inward current (192 ± 36 pA).

The co‐application of ZD 7288 and Ba2+ revealed a depolarizing sag in negative voltage deviations under current clamp conditions, or a large inward current with kinetics two to three times slower than those of Ih under voltage clamp conditions. This novel form of time‐dependent rectification was first apparent at potentials more negative than about −85 mV, was sensitive to 5 mm Cs+ (n= 4), and is termed Ih,slow. Ih,slow tail currents reversed between −65.3 and −56.6 mV (with potassium acetate electrodes, n= 3) or −57.6 and −50.3 mV (with KCl electrodes, n= 3).

Computer simulations confirmed that the pattern of anomalous rectification in thalamocortical neurones of the cat ventrobasal thalamus is mediated by the concerted action of Ih and a Ba2+ ‐sensitive current with properties similar to an inwardly rectifying K+ current (IKIR).
Phasic activity in the human erector spinae during repetitive hand movements
Authors: Zedka Milan , Prochazka Arthur
Published: September 2004

Phasic activity in the human back muscle erector spinae (ES) was studied during repetitive hand movements. The hand movements were elicited voluntarily by the subject or induced passively by the experimenter through a servomotor or through cyclical electrical stimulation of muscles acting about the wrist. The aim of the study was to determine whether the rhythmical activation of ES was of supraspinal, intersegmental or segmental origin.

When voluntary rhythmical hand movements were performed as fast as possible, cyclical ES EMG bursts occurred at the same frequency. This frequency was significantly higher than that reached when the task was to contract the back muscles as rapidly as possible. This suggests that the ES activity during the fast hand movements was not generated by direct commands descending to the ES muscles from the motor area of the cerebral cortex responsible for voluntary back muscle activation.

During imposed rhythmical hand movements, ES EMG bursts remained entrained to the hand movements, even when movement frequencies far exceeded those attainable voluntarily either for the hand or the back. This showed that ES EMG responses could be evoked by the hand movements even when these were not generated by descending neural commands. Two alternative mechanisms of ES activation were considered: (a) propriospinal transmission of afferent input entering the spinal cord from the upper extremity; (b) afferent input from ES and other trunk muscles, responding to local oscillations transmitted mechanically from the hand to the lower back.

Activation of ES via proprioceptive signals from the forearm was unlikely since (a) simultaneous electrical stimulation of wrist extensor and wrist flexor muscles did not result in repetitive ES EMG bursting; (b) cyclical vibration of the wrist extensors did not evoke ES EMG bursting; (c) when the forearm was constrained and the hand was moved passively, the lower trunk accelerations and cyclical ES EMG both occurred at a harmonic of the hand movement frequency.

We conclude that the repetitive ES EMG bursting during hand movements was probably due to a local segmental reflex rather than to descending commands. Remote mechanical oscillations of the trunk caused by hand movements evidently elicited proprioceptive reflexes in ES that presumably contributed to trunk stabilization.