Di-5-ANEQ(F)PTEA Offers Better Performance than Di-4-ANEQ(F)PTEA for In-Situ Cardiac Optical Mapping.

Cardiac optical mapping has traditionally been performed in ex-vivo, motion-arrested hearts. Recently, in-situ cardiac optical mapping has been made possible by both motion correction techniques and long-wavelength voltage sensitive dyes (VSDs). However, VSDs have been observed to wash out quickly from blood-perfused in-situ hearts.

Optical mapping of cardiac electromechanics in beating in vivo hearts.

Optical mapping has been widely used in the study of cardiac electrophysiology in motion-arrested, ex vivo heart preparations. Recent developments in motion artifact mitigation techniques have made it possible to optically map beating ex vivo hearts, enabling the study of cardiac electromechanics using optical mapping. However, the ex vivo setting imposes limitations on optical mapping such as altered metabolic states, oversimplified mechanical loads, and the absence of neurohormonal regulation.

Near-infrared voltage-sensitive dyes based on chromene donor.

Voltage-sensitive dyes (VSDs) are used to image electrical activity in cells and tissues with submillisecond time resolution. Most of these fast sensors are constructed from push-pull chromophores whose fluorescence spectra are modulated by the electric field across the cell membrane. It was found that the substitution of naphthalene with chromene produces a 60 to 80 nm red-shift in absorption and emission spectra while maintaining fluorescence quantum efficiency and voltage sensitivity.

Voltage imaging reveals the dynamic electrical signatures of human breast cancer cells.

Cancer cells feature a resting membrane potential (V) that is depolarized compared to normal cells, and express active ionic conductances, which factor directly in their pathophysiological behavior. Despite similarities to 'excitable' tissues, relatively little is known about cancer cell V dynamics. Here high-throughput, cellular-resolution V imaging reveals that V fluctuates dynamically in several breast cancer cell lines compared to non-cancerous MCF-10A cells.

Recent progress in optical voltage-sensor technology and applications to cardiac research: from single cells to whole hearts.

The first workshop on Novel Optics-based approaches for Cardiac Electrophysiology (NOtiCE) was held in Florence Italy in 2018. Here, we learned how optical approaches have shaped our basic understanding of cardiac electrophysiology and how new technologies and approaches are being developed and validated to advance the field. Several technologies are being developed that may one day allow for new clinical approaches for diagnosing cardiac disorders and possibly intervening to treat human patients.

Tethered Bichromophoric Fluorophore Quencher Voltage Sensitive Dyes.

Voltage sensitive dyes (VSDs) are used for in vitro drug screening and for imaging of patterns of electrical activity in tissue. Wide application of this technology depends on the availability of sensors with high sensitivity (percent change of fluorescence per 100 mV), high fluorescence quantum yield, and fast response kinetics. A promising approach uses a two-component system consisting of anionic membrane permeable quenchers with fluorophores labeling one side of the membrane; this produces voltage-dependent fluorescence quenching.

The stochastic nature of action potential backpropagation in apical tuft dendrites.

In cortical pyramidal neurons, backpropagating action potentials (bAPs) supply Ca to synaptic contacts on dendrites. To determine whether the efficacy of AP backpropagation into apical tuft dendrites is stable over time, we performed dendritic Ca and voltage imaging in rat brain slices. We found that the amplitude of bAP-Ca in apical tuft branches was unstable, given that it varied from trial to trial (termed "bAP-Ca flickering").

EPSPs Measured in Proximal Dendritic Spines of Cortical Pyramidal Neurons.

EPSPs occur when the neurotransmitter glutamate binds to postsynaptic receptors located on small pleomorphic membrane protrusions called dendritic spines. To transmit the synaptic signal, these potentials must travel through the spine neck and the dendritic tree to reach the soma. Due to their small size, the electrical behavior of spines and their ability to compartmentalize electrical signals has been very difficult to assess experimentally.

Characterization of voltage-sensitive dyes in living cells using two-photon excitation.

In this protocol, we describe the procedures we have developed to optimize the performance of voltage-sensitive dyes for recording changes in neuronal electrical activity. We emphasize our experience in finding the best dye conditions for recording backpropagating action potentials from individual dendritic spines in a neuron within a brain slice. We fully describe procedures for loading the dye through a patch pipette and for finding excitation and emission wavelengths for the best sensitivity of the fluorescence signal to membrane voltage.

Palette of fluorinated voltage-sensitive hemicyanine dyes.

Optical recording of membrane potential permits spatially resolved measurement of electrical activity in subcellular regions of single cells, which would be inaccessible to electrodes, and imaging of spatiotemporal patterns of action potential propagation in excitable tissues, such as the brain or heart. However, the available voltage-sensitive dyes (VSDs) are not always spectrally compatible with newly available optical technologies for sensing or manipulating the physiological state of a system. Here, we describe a series of 19 fluorinated VSDs based on the hemicyanine class of chromophores.

Single-voxel recording of voltage transients in dendritic spines.

We report sensitive recording of membrane potential in single dendritic spines in cortical neurons within a brain slice using two-photon excitation and a new, fluorinated, intracellularly loaded organic dye, di-2-AN(F)EPPTEA. With a two-photon excitation wavelength of 1060 nm, we achieve voltage sensitivity of >16% change in fluorescence per 100 mV. By targeting single spines in single-voxel recordings, we attain excellent single/noise quality, with back-propagating action potentials (bAPs) visible in single sweeps while recording at 10 kHz.

Quantitative assessment of the distributions of membrane conductances involved in action potential backpropagation along basal dendrites.

Basal dendrites of prefrontal cortical neurons receive strong synaptic drive from recurrent excitatory synaptic inputs. Synaptic integration within basal dendrites is therefore likely to play an important role in cortical information processing. Both synaptic integration and synaptic plasticity depend crucially on dendritic membrane excitability and the backpropagation of action potentials.

Roles of IA and morphology in action potential propagation in CA1 pyramidal cell dendrites.

Dendrites of CA1 pyramidal cells of the hippocampus, along with those of a wide range of other cell types, support active backpropagation of axonal action potentials. Consistent with previous work, recent experiments demonstrating that properties of synaptic plasticity are different for distal synapses, suggest an important functional role of bAPs, which are known to be prone to failure in distal locations. Using conductance-based models of CA1 pyramidal cells, we show that underlying "traveling wave attractors" control action potential propagation in the apical dendrites.

Increasing Ca2+ transients by broadening postsynaptic action potentials enhances timing-dependent synaptic depression.

Repeated induction of pre- and postsynaptic action potentials (APs) at a fixed time difference leads to long-term potentiation (LTP) or long-term depression (LTD) of the synapse, depending on the temporal order of pre- and postsynaptic activity. This phenomenon of spike-timing-dependent plasticity (STDP) is believed to arise by nonlinear processes that lead to larger calcium transients (and thus LTP) when presynaptic APs precede postsynaptic APs and smaller calcium transients (and thus LTD) when postsynaptic APs precede presynaptic APs. In contrast to predictions from such calcium-peak-detector models, we show that constitutively or artificially broadened APs in layer II/III pyramidal cells of entorhinal cortex (EC) lead to an increase in the dendritic calcium transient and shift the balance of STDP toward LTD.

Slow and fast inhibition and an H-current interact to create a theta rhythm in a model of CA1 interneuron network.

The oriens-lacunosum moleculare (O-LM) subtype of interneuron is a key component in the formation of the theta rhythm (8-12 Hz) in the hippocampus. It is known that the CA1 region of the hippocampus can produce theta rhythms in vitro with all ionotropic excitation blocked, but the mechanisms by which this rhythmicity happens were previously unknown. Here we present a model suggesting that individual O-LM cells, by themselves, are capable of producing a single-cell theta-frequency firing, but coupled O-LM cells are not capable of producing a coherent population theta.

Beyond two-cell networks: experimental measurement of neuronal responses to multiple synaptic inputs.

Oscillations of large populations of neurons are thought to be important in the normal functioning of the brain. We have used phase response curve (PRC) methods to characterize the dynamics of single neurons and predict population dynamics. Our past experimental work was limited to special circumstances (e.

Synchronization in hybrid neuronal networks of the hippocampal formation.

Understanding the mechanistic bases of neuronal synchronization is a current challenge in quantitative neuroscience. We studied this problem in two putative cellular pacemakers of the mammalian hippocampal theta rhythm: glutamatergic stellate cells (SCs) of the medial entorhinal cortex and GABAergic oriens-lacunosum-molecular (O-LM) interneurons of hippocampal region CA1. We used two experimental methods.

Synchronization of strongly coupled excitatory neurons: relating network behavior to biophysics.

Behavior of a network of neurons is closely tied to the properties of the individual neurons. We study this relationship in models of layer II stellate cells (SCs) of the medial entorhinal cortex. SCs are thought to contribute to the mammalian theta rhythm (4-12 Hz), and are notable for the slow ionic conductances that constrain them to fire at rates within this frequency range.