scholarly journals In vitro neuronal networks show bidirectional axonal conduction with antidromic action potentials effectively depolarizing the soma

2021 ◽  
Author(s):  
JC Mateus ◽  
CDF Lopes ◽  
M Aroso ◽  
AR Costa ◽  
A Gerós ◽  
...  
Keyword(s):  
Action Potential ◽  
Action Potentials ◽  
Neuronal Networks ◽  
Neuronal Cultures ◽  
Neuronal Function ◽  
Axonal Conduction ◽  
Technological Advances ◽  
Transduction Mechanisms ◽  
Or Gene

ABSTRACTRecent technological advances are revealing the complex physiology of the axon and challenging long-standing assumptions. Namely, while most action potential (AP) initiation occurs at the axon initial segment in central nervous system neurons, initiation in distal parts of the axon has been shown to occur in both physiological and pathological conditions. However, such ectopic action potential (EAP) activity has not been reported yet in studies using in vitro neuronal networks and its functional role, if exists, is still not clear. Here, we report the spontaneous occurrence of EAPs and effective antidromic conduction in hippocampal neuronal cultures. We also observe a significant fraction of bidirection axonal conduction in dorsal root ganglia neuronal cultures. We set out to investigate and characterize this antidromic propagation via a combination of microelectrode arrays, microfluidics, advanced data analysis and in silico studies. We show that EAPs and antidromic conduction can occur spontaneously, and also after distal axotomy or physiological changes in the axon biochemical environment. Importantly, EAPs may carry information (as orthodromic action potentials do) and can have a functional impact on the neuron, as they consistently depolarize the soma. Plasticity or gene transduction mechanisms triggered by soma depolarization can, therefore, be also affected by these antidromic action potentials/EAPs. Finally, we show that this bidirectional axonal conduction is asymmetrical, with antidromic conduction being slower than orthodromic. Via computational modeling, we show that the experimental difference can be explained by axonal morphology. Altogether, these findings have important implications for the study of neuronal function in vitro, reshaping completely our understanding on how information flows in neuronal cultures.

Bidirectional flow of action potentials in axons drives activity dynamics in neuronal cultures

2021 ◽  
Author(s):  
José Mateus ◽  
Cátia Lopes ◽  
Miguel Aroso ◽  
Ana Costa ◽  
Ana Geros ◽  
...  
Keyword(s):  
Network Activity ◽  
Neuronal Cultures ◽  
Axonal Conduction ◽  
Technological Advances ◽  
Transduction Mechanisms ◽  
Or Gene ◽  
Bidirectional Propagation ◽  
Bidirectional Flow ◽  
Activity Dynamics

Abstract Objective: Recent technological advances are revealing the complex physiology of the axon and challenging long-standing assumptions. Namely, while most action potential (AP) initiation occurs at the axon initial segment in central nervous system neurons, initiation in distal parts of the axon has been reported to occur in both physiological and pathological conditions. The functional role of these ectopic APs, if exists, is still not clear, nor its impact on network activity dynamics. Approach: Using an electrophysiology platform specifically designed for assessing axonal conduction we show here for the first time regular and effective bidirectional axonal conduction in hippocampal and dorsal root ganglia cultures. We investigate and characterize this bidirectional propagation both in physiological conditions and after distal axotomy. Main results: A significant fraction of APs are not coming from the canonical synapse-dendrite-soma signal flow, but instead from signals originating at the distal axon. Importantly, antidromic APs may carry information and can have a functional impact on the neuron, as they consistently depolarize the soma. Thus, plasticity or gene transduction mechanisms triggered by soma depolarization can also be affected by these antidromic APs. Conduction velocity is asymmetrical, with antidromic conduction being slower than orthodromic. Significance: Altogether these findings have important implications for the study of neuronal function in vitro, reshaping our understanding on how information flows in neuronal cultures.

scholarly journals Cell–cell coupling and DNA methylation abnormal phenotypes in the after-hours mice

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Federico Tinarelli ◽  
Elena Ivanova ◽  
Ilaria Colombi ◽  
Erica Barini ◽  
Edoardo Balzani ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Neuronal Networks ◽  
Molecular Mechanisms ◽  
Ex Vivo ◽  
Neuronal Cultures ◽  
Functional Impairments ◽  
After Hours ◽  
The Impact

Abstract Background DNA methylation has emerged as an important epigenetic regulator of brain processes, including circadian rhythms. However, how DNA methylation intervenes between environmental signals, such as light entrainment, and the transcriptional and translational molecular mechanisms of the cellular clock is currently unknown. Here, we studied the after-hours mice, which have a point mutation in the Fbxl3 gene and a lengthened circadian period. Methods In this study, we used a combination of in vivo, ex vivo and in vitro approaches. We measured retinal responses in Afh animals and we have run reduced representation bisulphite sequencing (RRBS), pyrosequencing and gene expression analysis in a variety of brain tissues ex vivo. In vitro, we used primary neuronal cultures combined to micro electrode array (MEA) technology and gene expression. Results We observed functional impairments in mutant neuronal networks, and a reduction in the retinal responses to light-dependent stimuli. We detected abnormalities in the expression of photoreceptive melanopsin (OPN4). Furthermore, we identified alterations in the DNA methylation pathways throughout the retinohypothalamic tract terminals and links between the transcription factor Rev-Erbα and Fbxl3. Conclusions The results of this study, primarily represent a contribution towards an understanding of electrophysiological and molecular phenotypic responses to external stimuli in the Afh model. Moreover, as DNA methylation has recently emerged as a new regulator of neuronal networks with important consequences for circadian behaviour, we discuss the impact of the Afh mutation on the epigenetic landscape of circadian biology.

Contribution of Nav1.8 Sodium Channels to Action Potential Electrogenesis in DRG Neurons

2001 ◽  
Vol 86 (2) ◽  
pp. 629-640 ◽  
Author(s):  
Muthukrishnan Renganathan ◽  
Theodore R. Cummins ◽  
Stephen G. Waxman
Keyword(s):  
Action Potential ◽  
Sodium Channels ◽  
Action Potentials ◽  
Input Resistance ◽  
Resting Membrane Potential ◽  
Drg Neurons ◽  
Significant Difference ◽  
Steady State Inactivation ◽  
Current Threshold

C-type dorsal root ganglion (DRG) neurons can generate tetrodotoxin-resistant (TTX-R) sodium-dependent action potentials. However, multiple sodium channels are expressed in these neurons, and the molecular identity of the TTX-R sodium channels that contribute to action potential production in these neurons has not been established. In this study, we used current-clamp recordings to compare action potential electrogenesis in Nav1.8 (+/+) and (−/−) small DRG neurons maintained for 2–8 h in vitro to examine the role of sodium channel Nav1.8 (α-SNS) in action potential electrogenesis. Although there was no significant difference in resting membrane potential, input resistance, current threshold, or voltage threshold in Nav1.8 (+/+) and (−/−) DRG neurons, there were significant differences in action potential electrogenesis. Most Nav1.8 (+/+) neurons generate all-or-none action potentials, whereas most of Nav1.8 (−/−) neurons produce smaller graded responses. The peak of the response was significantly reduced in Nav1.8 (−/−) neurons [31.5 ± 2.2 (SE) mV] compared with Nav1.8 (+/+) neurons (55.0 ± 4.3 mV). The maximum rise slope was 84.7 ± 11.2 mV/ms in Nav1.8 (+/+) neurons, significantly faster than in Nav1.8 (−/−) neurons where it was 47.2 ± 1.3 mV/ms. Calculations based on the action potential overshoot in Nav1.8 (+/+) and (−/−) neurons, following blockade of Ca2+ currents, indicate that Nav1.8 contributes a substantial fraction (80–90%) of the inward membrane current that flows during the rising phase of the action potential. We found that fast TTX-sensitive Na+ channels can produce all-or-none action potentials in some Nav1.8 (−/−) neurons but, presumably as a result of steady-state inactivation of these channels, electrogenesis in Nav1.8 (−/−) neurons is more sensitive to membrane depolarization than in Nav1.8 (+/+) neurons, and, in the absence of Nav1.8, is attenuated with even modest depolarization. These observations indicate that Nav1.8 contributes substantially to action potential electrogenesis in C-type DRG neurons.

scholarly journals Versatile live-cell activity analysis platform for characterization of neuronal dynamics at single-cell and network level

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Xinyue Yuan ◽  
Manuel Schröter ◽  
Marie Engelene J. Obien ◽  
Michele Fiscella ◽  
Wei Gong ◽  
...  
Keyword(s):  
Neuronal Networks ◽  
Microelectrode Array ◽  
Cell Activity ◽  
Live Cell ◽  
High Signal ◽  
Neuronal Cultures ◽  
Label Free ◽  
Fluorescent Indicators ◽  
Analysis Platform

AbstractChronic imaging of neuronal networks in vitro has provided fundamental insights into mechanisms underlying neuronal function. Current labeling and optical imaging methods, however, cannot be used for continuous and long-term recordings of the dynamics and evolution of neuronal networks, as fluorescent indicators can cause phototoxicity. Here, we introduce a versatile platform for label-free, comprehensive and detailed electrophysiological live-cell imaging of various neurogenic cells and tissues over extended time scales. We report on a dual-mode high-density microelectrode array, which can simultaneously record in (i) full-frame mode with 19,584 recording sites and (ii) high-signal-to-noise mode with 246 channels. We set out to demonstrate the capabilities of this platform with recordings from primary and iPSC-derived neuronal cultures and tissue preparations over several weeks, providing detailed morpho-electrical phenotypic parameters at subcellular, cellular and network level. Moreover, we develop reliable analysis tools, which drastically increase the throughput to infer axonal morphology and conduction speed.

Transduction mechanisms in airway sensory nerves

2006 ◽  
Vol 101 (3) ◽  
pp. 950-959 ◽  
Author(s):  
Thomas Taylor-Clark ◽  
Bradley J. Undem
Keyword(s):  
Action Potential ◽  
Action Potentials ◽  
Membrane Depolarization ◽  
Sensory Nerves ◽  
Metabotropic Receptors ◽  
Metabotropic Receptor ◽  
Transduction Mechanisms ◽  
The Central Nervous System ◽  
Action Potential Initiation ◽  
Potential Initiation

The induction of action potentials in airway sensory nerves relies on events leading to the opening of cation channels in the nerve terminal membrane and subsequent membrane depolarization. If the membrane depolarization is of sufficient rate and amplitude, action potential initiation will occur. The action potentials are then conducted to the central nervous system, leading to the initiation of various sensations and cardiorespiratory reflexes. Triggering events in airway sensory nerves include mechanical perturbation, inflammatory mediators, pH, temperature, and osmolarity acting through a variety of ionotropic and metabotropic receptors. Action potential initiation can be modulated (positively or negatively) through independent mechanisms caused mainly by autacoids and other metabotropic receptor ligands. Finally, gene expression of sensory nerves can be altered in adult mammals. This neuroplasticity can change the function of sensory nerves and likely involve both neurotrophin and use-dependent mechanisms. Here we provide a brief overview of some of the transduction mechanisms underlying these events.

scholarly journals Two Components of the Cardiac Action Potential

1969 ◽  
Vol 54 (5) ◽  
pp. 607-635 ◽  
Author(s):  
Antonio Paes de Carvalho ◽  
Brian Francis Hoffman ◽  
Marilene de Paula Carvalho
Keyword(s):  
Action Potential ◽  
Action Potentials ◽  
Time Course ◽  
Slow Component ◽  
Slow Inward Current ◽  
Equilibrium Potential ◽  
Slow Phase ◽  
Positive Equilibrium ◽  
Cardiac Action

Transmembrane potentials recorded from the rabbit heart in vitro were displayed as voltage against time (V, t display), and dV/dt against voltage (V, V or phase-plane display). Acetylcholine was applied to the recording site by means of a hydraulic system. Results showed that (a) differences in time course of action potential upstroke can be explained in terms of the relative magnitude of fast and slow phases of depolarization; (b) acetylcholine is capable of depressing the slow phase of depolarization as well as the plateau of the action potential; and (c) action potentials from nodal (SA and AV) cells seem to lack the initial fast phase. These results were construed to support a two-component hypothesis for cardiac electrogenesis. The hypothesis states that cardiac action potentials are composed of two distinct and physiologically separable "components" which result from discrete mechanisms. An initial fast component is a sodium spike similar to that of squid nerve. The slow component, which accounts for both a slow depolarization during phase 0 and the plateau, probably is dependent on the properties of a slow inward current having a positive equilibrium potential, coupled to a decrease in the resting potassium conductance. According to the hypothesis, SA and AV nodal action potentials are due entirely or almost entirely to the slow component and can therefore be expected to exhibit unique electrophysiological and pharmacological properties.

Transmembrane potential and force of contraction of the normal and potassium-deficient human atrial tissue in vitro

1969 ◽  
Vol 47 (12) ◽  
pp. 1015-1024 ◽  
Author(s):  
Kailash Prasad
Keyword(s):  
Action Potential ◽  
Action Potentials ◽  
Terminal Phase ◽  
Free Solution ◽  
Force Of Contraction ◽  
Pacemaker Cells ◽  
Atrial Tissue ◽  
Action Potential Plateau ◽  
Potential Plateau

The characteristics of the simultaneously recorded action potential (AP) and contraction of isolated atrial tissue of human heart were studied in normal and in potassium-free solutions. Two types of action potentials associated with characteristic contractions were observed. Pacemaker types of action potentials with two humps in the plateau were observed in spontaneously beating atria and they were associated with triple contractions. A non-pacemaker type of action potential was found in quiescent fibers when the preparation was driven electrically; this resulted in single peak contraction. The sizes of the resting and action potentials of pacemaker cells were lower, while those of action potential duration in all types of cells were higher than those reported in other mammals. When spontaneously beating atrial pieces were exposed to a potassium-free solution there was a shortening of the action potential plateau and lengthening of the terminal phase of repolarization associated with the development of the first hump into a slow spike. The shortening of the action potential plateau associated with an increase in the force of contraction was also observed in electrically stimulated muscles in KCl-free solution.

Axon terminal hyperexcitability associated with epileptogenesis in vitro. I. Origin of ectopic spikes

1993 ◽  
Vol 70 (3) ◽  
pp. 961-975 ◽  
Author(s):  
S. F. Stasheff ◽  
M. Hines ◽  
W. A. Wilson
Keyword(s):  
Action Potential ◽  
Action Potentials ◽  
Pyramidal Neurons ◽  
Model Neuron ◽  
Extracellular Recording ◽  
Pyramidal Cells ◽  
Initiation Site ◽  
Electrographic Seizures ◽  
Ca3 Pyramidal Cells

1. Intracellular and extracellular recording techniques were used to study the increase in ectopic (i.e., nonsomatic) action-potential generation occurring among CA3 pyramidal cells during the kindling-like induction of electrographic seizures (EGSs) in this subpopulation of the hippocampal slice. Kindling-like stimulus trains (60 Hz, 2 s) were delivered to s. radiatum of CA3 at 10-min intervals. As EGSs developed, the frequency of ectopic firing increased markedly (by 10.33 +/- 3.29 spikes/min, mean +/- SE, P << 0.01). Several methods were applied to determine the initiation site for these action potentials within the cell (axons vs. dendrites). 2. Collision tests were conducted between known antidromic and orthodromic action potentials in CA3 cells to determine the critical period, c, for collision. Attempts were then made to collide ectopic spikes with known antidromic action potentials. At intervals less than c, ectopic spikes failed to collide with antidromic ones, in 5 of 10 cases. In these cells, this clearly indicates that the ectopic spikes were themselves of axonal origin. In the remaining five cases, ectopic spikes collided with antidromic action potentials at intervals approximately equal to c, most likely because of interactions within the complex system of recurrent axon collaterals in CA3. 3. Action potentials of CA3 pyramidal cells were simulated with the use of a compartmental computer model, NEURON. These simulations were based on prior models of CA3 pyramidal neurons and of the motoneuron action potential. Simulated action potentials generated in axonal compartments possessed a prominent inflection on their rising phase (IS-SD break), which was difficult to appreciate in those spikes generated in somatic or dendritic compartments. 4. An analysis of action potentials recorded experimentally from CA3 pyramidal cells also showed that antidromic spikes possess a prominent IS-SD break that is not present in orthodromic spikes. In addition to identified antidromic action potentials, ectopic spikes also possess such an inflection. Together with the predictions of computer simulations, this analysis also indicates that ectopic spikes originate in the axons of CA3 cells. 5. Tetrodotoxin (TTX, 50 microM) was locally applied by pressure injection while monitoring ectopic spike activity. Localized application of TTX to regions of the slice that could include the axons but not the dendrites of recorded cells abolished or markedly reduced the frequency of ectopic spikes (n = 5), further confirming the hypothesis that these action potentials arise from CA3 axons.(ABSTRACT TRUNCATED AT 400 WORDS)

Properties of Action-Potential Initiation in Neocortical Pyramidal Cells: Evidence From Whole Cell Axon Recordings

2007 ◽  
Vol 97 (1) ◽  
pp. 746-760 ◽  
Author(s):  
Yousheng Shu ◽  
Alvaro Duque ◽  
Yuguo Yu ◽  
Bilal Haider ◽  
David A. McCormick
Keyword(s):  
Action Potential ◽  
Action Potentials ◽  
Initial Segment ◽  
Pyramidal Cells ◽  
Synaptic Activity ◽  
Up States ◽  
Action Potential Initiation ◽  
Potential Initiation

Cortical pyramidal cells are constantly bombarded by synaptic activity, much of which arises from other cortical neurons, both in normal conditions and during epileptic seizures. The action potentials generated by barrages of synaptic activity may exhibit a variable site of origin. Here we performed simultaneous whole cell recordings from the soma and axon or soma and apical dendrite of layer 5 pyramidal neurons during normal recurrent network activity (up states), the intrasomatic or intradendritic injection of artificial synaptic barrages, and during epileptiform discharges in vitro. We demonstrate that under all of these conditions, the real or artificial synaptic bombardments propagate through the dendrosomatic-axonal arbor and consistently initiate action potentials in the axon initial segment that then propagate to other parts of the cell. Action potentials recorded intracellularly in vivo during up states and in response to visual stimulation exhibit properties indicating that they are typically initiated in the axon. Intracortical axons were particularly well suited to faithfully follow the generation of action potentials by the axon initial segment. Action-potential generation was more reliable in the distal axon than at the soma during epileptiform activity. These results indicate that the axon is the preferred site of action-potential initiation in cortical pyramidal cells, both in vivo and in vitro, with state-dependent back propagation through the somatic and dendritic compartments.

scholarly journals Nicotine suppresses hyperexcitability of colonic sensory neurons and visceral hypersensivity in mouse model of colonic inflammation

2012 ◽  
Vol 302 (7) ◽  
pp. G740-G747 ◽  
Author(s):  
Galya R. Abdrakhmanova ◽  
Minho Kang ◽  
M. Imad Damaj ◽  
Hamid I. Akbarali
Keyword(s):  
Mouse Model ◽  
Action Potential ◽  
Action Potentials ◽  
Drg Neurons ◽  
Action Potential Firing ◽  
Colonic Inflammation ◽  
Single Action ◽  
Nicotine Administration ◽  
Potential Firing

Recently, we reported that nicotine in vitro at a low 1-μM concentration suppresses hyperexcitability of colonic dorsal root ganglia (DRG; L1-L2) neurons in the dextran sodium sulfate (DSS)-induced mouse model of acute colonic inflammation ( 1 ). Here we show that multiple action potential firing in colonic DRG neurons persisted at least for 3 wk post-DSS administration while the inflammatory signs were diminished. Similar to that in DSS-induced acute colitis, bath-applied nicotine (1 μM) gradually reduced regenerative multiple-spike action potentials in colonic DRG neurons to a single action potential in 3 wk post-DSS neurons. Nicotine (1 μM) shifted the activation curve for tetrodotoxin (TTX)-resistant sodium currents in inflamed colonic DRG neurons (voltage of half-activation changed from −37 to −32 mV) but did not affect TTX-sensitive currents in control colonic DRG neurons. Further, subcutaneous nicotine administration (2 mg/kg b.i.d.) in DSS-treated C57Bl/J6 male mice resulted in suppression of hyperexcitability of colonic DRG (L1-L2) neurons and the number of abdominal constrictions in response to intraperitoneal injection of 0.6% acetic acid. Collectively, the data suggest that neuronal nicotinic acetylcholine receptor-mediated suppression of hyperexcitability of colonic DRG neurons attenuates reduction of visceral hypersensitivity in DSS mouse model of colonic inflammation.

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