scholarly journals Imaging action potential in single mammalian neurons by tracking the accompanying sub-nanometer mechanical motion

2017 ◽  
Author(s):  
Yunze Yang ◽  
Xian-Wei Liu ◽  
Hui Yu ◽  
Yan Guan ◽  
Shaopeng Wang ◽  
...  
Keyword(s):  
Detection Limit ◽  
Patch Clamp ◽  
Action Potential ◽  
Fluorescence Detection ◽  
Action Potentials ◽  
Single Cells ◽  
Detection Methods ◽  
Imaging Method ◽  
Label Free ◽  
Mechanical Motion

AbstractAction potentials in neurons have been studied traditionally by the patch clamp and more recently by the fluorescence detection methods. Here we describe a label-free optical imaging method that can measure mechanical motion in single cells with sub-nanometer detection limit and sub-millisecond temporal resolution. Using the method, we have observed sub-nanometer mechanical motion accompanying the action potential in single mammalian neurons. The shape and width of the transient displacement are similar to those of the electrically recorded action potential, but the amplitude varies from neuron to neuron, and from one region of a neuron to another, ranging from 0.2 - 0.4 nm. The work indicates that action potentials may be studied non-invasively in single mammalian neurons by label-free imaging of the accompanying subnanometer mechanical motion.

scholarly journals Propagation of Action Potentials From the Soma to Individual Dendrite of Cultured Rat Amacrine Cells Is Regulated by Local GABA Input

2002 ◽  
Vol 87 (6) ◽  
pp. 2858-2866 ◽  
Author(s):  
Yoshitake Yamada ◽  
Amane Koizumi ◽  
Eisuke Iwasaki ◽  
Shu-Ichi Watanabe ◽  
Akimichi Kaneko
Keyword(s):  
Patch Clamp ◽  
Action Potential ◽  
Lateral Inhibition ◽  
Action Potentials ◽  
Amacrine Cell ◽  
Amacrine Cells ◽  
Inner Plexiform Layer ◽  
Whole Cell ◽  
Cell Patch Clamp ◽  
Whole Cell Patch Clamp

Retinal amacrine cells are interneurons that make lateral and vertical connections in the inner plexiform layer of the retina. Amacrine cells do not possess a long axon, and this morphological feature is the origin of their naming. Their dendrites function as both presynaptic and postsynaptic sites. Half of all amacrine cells are GABAergic inhibitory neurons that mediate lateral inhibition, and their light-evoked response consists of graded voltage changes and regenerative action potentials. There is evidence that the amount of neurotransmitter release from presynaptic sites is increased by spike propagation into the dendrite. Thus understanding of how action potentials propagate in dendrites is important to elucidating the extent and strength of lateral inhibition. In the present study, we used the dual whole cell patch-clamp technique on the soma and the dendrite of cultured rat amacrine cells and directly demonstrated that the action potentials propagate into the dendrites. The action potential in the dendrite was TTX sensitive and was affected by the local membrane potential of the dendrite. Propagation of the action potential was suppressed by local application of GABA to the dendrite. Dual dendrite whole cell patch-clamp recordings showed that GABA suppresses the propagation of action potentials in one dendrite of an amacrine cell, while the action potentials propagate in the other dendrites. It is likely that the action potentials in the dendrites are susceptible to various external factors resulting in the nonuniform propagation of the action potential from the soma of an amacrine cell.

Changes in cytosolic calcium monitored by inward currents during action potentials in guinea-pig ventricular cells

1989 ◽  
Vol 238 (1291) ◽  
pp. 171-188 ◽  
Keyword(s):  
Guinea Pig ◽  
Action Potential ◽  
Voltage Clamp ◽  
Action Potentials ◽  
Single Cells ◽  
Calcium Transient ◽  
Cytosolic Calcium ◽  
Time To Peak ◽  
Ventricular Cells ◽  
In Cells

Action potentials were recorded from single cells isolated from guinea-pig ventricular muscle. Contraction was measured with an optical technique. Tail currents thought to be activated by cytosolic calcium were recorded when action potentials were interrupted by application of a voltage-clamp. A family of tail currents was recorded by interrupting the action potential at various times after the upstroke. The envelope of tail current amplitudes was taken as an index of changes in cytosoli calcium. Con­sistent with this interpretation, tail currents were negligible following intracellular loading with the calcium chelator BAPTA to suppress calcium transients. The cytosolic calcium transient estimated from the envelope of tails reached a peak approximately 50 ms after the upstroke of the action potential, and fell close to diastolic levels before repolarization was com­plete; 10 mM caffeine delayed the time to peak contraction, and caused a prolongation of the cytosolic calcium transient estimated from the envelope of tail currents. Caffeine also induced the appearance of a distinct late plateau phase of the action potential. Intracellular BAPTA suppressed the late plateau, contraction and tail currents in cells exposed to caffeine. Exposure to caffeine increased the time constant for decay of tail currents (from approximately 35 to 70 ms). When action potentials were greatly abbreviated by interruption with a voltage-clamp, a pro­gressive decline occurred in the subsequent three contractions and tail currents. There was a progressive reversal of these effects over four responses when the full action potential duration was restored. None of these effects was observed in cells exposed to caffeine. Calcium-activated tail currents appear to be a useful qualitative index of changes in cytosolic calcium. The observations are consistent with the suggestion that cytosolic calcium is reduced during the plateau by a combination of calcium extrusion through Na–Ca exchange and calcium uptake into caffeine-sensitive stores. It also appears that reduction of stores loading during abbreviated action potentials reduces subsequent contraction in cells not exposed to caffeine.

scholarly journals Diverse Ionic Currents and Electrical Activity of Cultured Myenteric Neurons From the Guinea Pig Proximal Colon

2000 ◽  
Vol 83 (3) ◽  
pp. 1253-1263 ◽  
Author(s):  
Fivos Vogalis ◽  
Kirk Hillsley ◽  
Terence K. Smith
Keyword(s):  
Patch Clamp ◽  
Guinea Pig ◽  
Action Potential ◽  
Action Potentials ◽  
Outward Current ◽  
Proximal Colon ◽  
Equilibrium Potential ◽  
Transient Outward Current ◽  
Myenteric Neurons ◽  
Fast Transient

The aim of this study was to perform a patch-clamp analysis of myenteric neurons from the guinea pig proximal colon. Neurons were enzymatically dispersed, cultured for 2–7 days, and recorded from using whole cell patch clamp. The majority of cells fired phasically, whereas about one-quarter of the neurons fired in a tonic manner. Neurons were divided into three types based on the currents activated. The majority of tonically firing neurons lacked an A-type current, but generated a large fast transient outward current that was associated with the rapid repolarizing phase of an action potential. The fast transient outward current was dependent on calcium entry and was blocked by tetraethylammonium. Cells that expressed both an A-type current and a fast transient outward current were mostly phasic. Depolarization of these cells to suprathreshold potentials from less than −60 mV failed to trigger action potentials, or action potentials were only triggered after a delay of >50 ms. However, depolarizations from more positive potentials triggered action potentials with minimal latency. Neurons that expressed neither the A-type current or the fast transient outward current were all phasic. Sixteen percent of neurons were similar to AH/type II neurons in that they generated a prolonged afterhyperpolarization following an action potential. The current underlying the prolonged afterhyperpolarization showed weak inward rectification and had a reversal potential near the potassium equilibrium potential. Thus cultured isolated myenteric neurons of the guinea pig proximal colon retain many of the diverse properties of intact neurons. This preparation is suitable for further biophysical and molecular characterization of channels expressed in colonic myenteric neurons.

Isolated myocytes from adult canine left ventricle: Ca2+ tolerance, electrophysiology, and ultrastructure

1983 ◽  
Vol 245 (5) ◽  
pp. H830-H839 ◽  
Author(s):  
K. Hewett ◽  
M. J. Legato ◽  
P. Danilo ◽  
R. B. Robinson
Keyword(s):  
Left Ventricle ◽  
Action Potential ◽  
Action Potentials ◽  
Single Cells ◽  
Resting Potential ◽  
High Yield ◽  
Intact Cells ◽  
Homogeneous Population ◽  
Ventricular Cells ◽  
Isolated Myocyte

We have developed a method for isolating single cardiac muscle cells in high yield (greater than 5 X 10(7) cells) from the canine left ventricle. Most of the myocytes are single cells with ultrastructural detail indistinguishable from intact ventricular myocardium, and more than 50% of the isolated cells remain elongated for at least 7 h in 0.5 mM calcium. Electrophysiological studies demonstrate that external potassium has a strong influence on repolarization in the isolated ventricular cells. Action potentials in [K+]o = 3.78 mM exhibit a positive over-shoot (greater than zero potential), but repolarization often arrests at congruent to -35 mV unless driven to more negative potentials by hyperpolarizing current. This phenomenon of two levels of resting potential is not observed at [K+]o = 5.78 mM. At the higher potassium concentration, values for maximum diastolic potential, amplitude, maximum rate of rise of phase 0, and action potential duration all are similar to those of intact ventricular muscle. However, the potential at the peak of the action potential plateau (phase 2) in the isolated myocyte is considerably more negative than that of intact myocardium. In addition, there is a conspicuous notch between phases 1 and 2 of the action potential in the isolated myocyte, whereas the notch is small or absent in intact myocardial action potentials. In summary, our method results in a preparation of stable, ultrastructurally and electrophysiologically intact cells, which should prove useful in studies requiring a large and homogeneous population of myocardial cells.

L-364,373 (R-L3) enantiomers have opposite modulating effects on IKs in mammalian ventricular myocytes

2013 ◽  
Vol 91 (8) ◽  
pp. 586-592 ◽  
Author(s):  
Claudia Corici ◽  
Zsófia Kohajda ◽  
Attila Kristóf ◽  
András Horváth ◽  
László Virág ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Guinea Pig ◽  
Action Potential ◽  
Right Ventricular ◽  
Action Potential Duration ◽  
Action Potentials ◽  
Ventricular Arrhythmias ◽  
Ventricular Myocytes ◽  
Delayed Rectifier ◽  
Whole Cell Patch Clamp

Activators of the slow delayed rectifier K+ current (IKs) have been suggested as promising tools for suppressing ventricular arrhythmias due to prolongation of repolarization. Recently, L-364,373 (R-L3) was nominated to activate IKs in myocytes from several species; however, in some studies, it failed to activate IKs. One later study suggested opposite modulating effects from the R-L3 enantiomers as a possible explanation for this discrepancy. Therefore, we analyzed the effect of the RL-3 enantiomers on IKs in ventricular mammalian myocytes, by applying standard microelectrode and whole-cell patch-clamp techniques at 37 °C. We synthesized 2 substances, ZS_1270B (right) and ZS_1271B (left), the 2 enantiomers of R-L3. In rabbit myocytes, ZS_1270B enhanced the IKs tail current by approximately 30%, whereas ZS_1271B reduced IKs tails by 45%. In guinea pig right ventricular preparations, ZS_1270B shortened APD90 (action potential duration measured at 90% repolarization) by 12%, whereas ZS_1271B lengthened it by approximately 15%. We concluded that R-L3 enantiomers in the same concentration range indeed have opposite modulating effects on IKs, which may explain why the racemic drug R-L3 previously failed to activate IKs. ZS_1270B is a potent IKs activator, therefore, this substance is appropriate to test whether IKs activators are ideal tools to suppress ventricular arrhythmias originating from prolongation of action potentials.

Ionic currents during action potentials in mammalian skeletal muscle fibers analyzed with loose patch clamp

1994 ◽  
Vol 267 (6) ◽  
pp. C1699-C1706 ◽  
Author(s):  
H. Wolters ◽  
W. Wallinga ◽  
D. L. Ypey ◽  
H. B. Boom
Keyword(s):  
Patch Clamp ◽  
Action Potential ◽  
Action Potentials ◽  
Potassium Current ◽  
Sodium Current ◽  
Delayed Rectifier ◽  
Mammalian Skeletal Muscle ◽  
Action Potential Generation ◽  
Potential Generation ◽  
Patch Clamp Technique

The loose patch-clamp technique was applied to analyze transmembrane currents during propagating action potentials in superficial fibers of musculi extensor digitorum longus of the mouse in vitro. Experimentally three components were identified in the transmembrane current: 1) a capacitive, 2) an inward sodium, and 3) an outward potassium current. Other components were negligible. The capacitive current was similar in shape to the first derivative of the intracellularly measured action potential. Tetrodotoxin, tetraethylammonium, and 4-aminopyridine, applied in the pipette, were used to identify the contribution in the current by sodium and potassium ions. With extracellularly applied depolarization steps only a sodium current was observed, not a potassium current. Occasionally found outward currents were artifactual. The behaviour of delayed rectifier potassium channels in muscle fiber membranes is discussed in the light of these unexpected findings. We conclude that potassium channel activity contributing to and measured during action potential generation is in some way inaccessible to loose patch extracellular voltage-clamp stimulation and that loose patch action current recording is a useful noninvasive method to analyze membrane conductances involved in action potential generation.

scholarly journals Label-Free Bacterial Imaging with Deep-UV-Laser-Induced Native Fluorescence

2010 ◽  
Vol 76 (21) ◽  
pp. 7231-7237 ◽  
Author(s):  
Rohit Bhartia ◽  
Everett C. Salas ◽  
William F. Hug ◽  
Ray D. Reid ◽  
Arthur L. Lane ◽  
...  
Keyword(s):  
Fluorescent Dyes ◽  
Spatial Scales ◽  
Single Cells ◽  
Deep Ocean ◽  
Imaging Method ◽  
Label Free ◽  
Deep Biosphere ◽  
Native Fluorescence ◽  
Deep Uv

ABSTRACT We introduce a near-real-time optical imaging method that works via the detection of the intrinsic fluorescence of life forms upon excitation by deep-UV (DUV) illumination. A DUV (<250-nm) source enables the detection of microbes in their native state on natural materials, avoiding background autofluorescence and without the need for fluorescent dyes or tags. We demonstrate that DUV-laser-induced native fluorescence can detect bacteria on opaque surfaces at spatial scales ranging from tens of centimeters to micrometers and from communities to single cells. Given exposure times of 100 μs and low excitation intensities, this technique enables rapid imaging of bacterial communities and cells without irreversible sample alteration or destruction. We also demonstrate the first noninvasive detection of bacteria on in situ-incubated environmental experimental samples from the deep ocean (Lo'ihi Seamount), showing the use of DUV native fluorescence for in situ detection in the deep biosphere and other nutrient-limited environments.

scholarly journals How neurons move during action potentials

2019 ◽  
Author(s):  
Tong Ling ◽  
Kevin C. Boyle ◽  
Valentina Zuckerman ◽  
Thomas Flores ◽  
Charu Ramakrishnan ◽  
...  
Keyword(s):  
Action Potential ◽  
Action Potentials ◽  
Time Course ◽  
Time Derivative ◽  
Optical Signal ◽  
Electrical Signal ◽  
Phase Changes ◽  
Phase Imaging ◽  
Label Free ◽  
Full Field

AbstractNeurons undergo nanometer-scale deformations during action potentials, and the underlying mechanism has been actively debated for decades. Previous observations were limited to a single spot or the cell boundary, while movement across the entire neuron during the action potential remained unclear.We report full-field imaging of cellular deformations accompanying the action potential in mammalian neuron somas (−1.8nm~1.3nm) and neurites (−0.7nm~0.9nm), using fast quantitative phase imaging with a temporal resolution of 0.1ms and an optical pathlength sensitivity of <4pm per pixel. Spike-triggered average, synchronized to electrical recording, demonstrates that the time course of the optical phase changes matches the dynamics of the electrical signal, with the optical signal revealing the intracellular potential rather than its time derivative detected via extracellular electrodes. Using 3D cellular morphology extracted via confocal microscopy, we demonstrate that the voltage-dependent changes in the membrane tension induced by ionic repulsion can explain the magnitude, time course and spatial features of the phase imaging. Our full-field observations of the spike-induced deformations in mammalian neurons opens the door to non-invasive label-free imaging of neural signaling.

Effect of isoprenaline, carbachol, and Cs+ on Na+ activity and pacemaker potential in rabbit SA node cells

1999 ◽  
Vol 276 (1) ◽  
pp. H205-H214 ◽  
Author(s):  
Ho S. Choi ◽  
Dai Y. Wang ◽  
Denis Noble ◽  
Chin O. Lee
Keyword(s):  
Action Potential ◽  
Action Potentials ◽  
Single Cells ◽  
Sa Node ◽  
Spontaneous Action ◽  
Sodium Binding ◽  
Potential Cycle ◽  
Spontaneous Action Potentials ◽  
Potential Rate ◽  
Zd 7288

Effects of isoprenaline, carbachol, and Cs+on intracellular Na+ activity ([Formula: see text]) and spontaneous action potentials were studied in multicellular and single cell preparations isolated from rabbit sinoatrial (SA) nodes.[Formula: see text] was measured with double-barreled Na+-selective microelectrodes and the fluorescent Na+-indicator sodium-binding benzofuran isophthalate (SBFI). In spontaneously beating cells,[Formula: see text] measured with Na+-selective microelectrodes and SBFI were 4.5 ± 1.2 mM (means ± SD, n = 21) in multicellular preparations and 4.0 ± 1.1 mM ( n = 16) in single cells, respectively. Measurements of[Formula: see text] with microelectrodes showed that isoprenaline increased[Formula: see text] from 4.7 ± 1.2 to 5.5 ± 1.6 mM ( n = 16, P < 0.01) and shortened the action potential cycle length (ACL) from 338 ± 46 to 269 ± 35 ms ( n = 16, P < 0.01). However, increasing the action potential rate by pacing produced a much smaller increase in[Formula: see text]. Changes in[Formula: see text] and ACL produced by isoprenaline were blocked by Cs+. The selective hyperpolarization-activated inward current ( I f) blocker ZD-7288 decreased [Formula: see text] from 5.2 ± 1.0 to 4.6 ± 1.3 mM ( n = 4, P < 0.01) and prolonged ACL from 394 ± 20 to 553 ± 68 ms ( n = 4, P < 0.01). The I f blocker substantially inhibited the increase in[Formula: see text] produced by isoprenaline. Carbachol and Cs+ decreased[Formula: see text] from 4.6 ± 1.4 to 3.9 ± 1.2 mM ( n = 15, P < 0.01) and from 4.9 ± 1.0 to 3.9 ± 1.3 mM ( n = 18, P < 0.01), respectively. In addition, carbachol and Cs+prolonged ACL from 345 ± 44 to 587 ± 100 ms ( n = 15, P < 0.01) and from 353 ± 30 to 464 ± 87 ms ( n = 18, P < 0.01), respectively. However, carbachol and Cs+ almost did not change [Formula: see text] when SA node cells became quiescent in a 25.4 mM extracellular K+ concentration. The results suggest that isoprenaline, ZD-7288, carbachol, or Cs+ might have changed[Formula: see text] and action potential rate by possibly stimulating or inhibiting I f carried by Na+. Measurements of[Formula: see text] with SBFI showed that isoprenaline, carbachol, and Cs+produced [Formula: see text] changes that were similar to those measured with the microelectrodes.

Sign in / Sign up

Export Citation Format

Share Document