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P Fromherz

Publications and source records attributed to P Fromherz.

At least 19 recordsLinked to original sources
PubMed1999

Transistor probes local potassium conductances in the adhesion region of cultured rat hippocampal neurons.

Adhesion interactions of neurons in a tissue may affect the ion conductance of the plasma membrane, inducing selective localization and modulation of channels. We studied the adhesion region of cultured neurons from rat hippocampus as a defined model where such effects could be observed electrophysiologically, taking advantage of extracellular recording by a transistor integrated in the substrate. We observed the K(+) current through the region of soma adhesion under voltage-clamp and compared it with the current through the whole cell. We found that the specific A-type conductance was depleted, even completely, in the region of adhesion, whereas the specific K-type conductance was enhanced up to a factor of 12. The electrophysiological approach opens a new way to investigate targeting of ion channels in the cell membrane as a function of adhesion processes.

Animals↗
PubMed1999

Extracellular recording with transistors and the distribution of ionic conductances in a cell membrane.

Intracellular voltage transients of cultured cells are recorded by transistors and other planar electrodes as local extracellular voltages. The theoretical relationship between extra- and intracellular voltage is investigated with a two-compartment circuit using the approximation of a fast, weak and small cell-silicon junction. It is shown that extracellular recording relies on the difference of specific ionic conductances in the attached and free regions of the cell membrane. The result rationalizes various observations with neuron transistors. It guides the optimization of extracellular recording and the development of cell-based chemical sensors.

Biophysical Phenomena↗
PubMed1999

Ca2+ activation of hSlo K+ channel is suppressed by N-terminal GFP tag.

The human slow poke (hSlo) K+ channel was tagged with GFP (green fluorescent protein) at the N-terminus of its alpha-subunit. The fusion protein was expressed transiently in HEK293 cells; it formed functional voltage-gated channels as shown by whole cell patch-clamp measurements. However, the tag lowered the voltage dependence of gating and it suppressed the typical left-shift of gating by intracellular binding of Ca2+. The location of the GFP-tagged N-terminus was confirmed to be on the extracellular side by application of a monoclonal antibody to nonpermeabilized cells. Structural interpretations of the effects are discussed.

Calcium↗
PubMed1998

Neuron-silicon junction with voltage-gated ionic currents.

We recorded the signals of firing Retzius neurons from Hirudo medicinalis by field-effect transistors. The axon stump of dissociated cells was attached to an open gate coated with concanavalin A. We observed a new type of neuron-transistor coupling: the extracellular voltage transients beneath the neuron were dominated by a negative peak during the rising phase of the action potential with a weaker positive transient in the falling phase. The biphasic response was opposite to the signal of capacitive coupling. We simulated the junction on the basis of the Hodgkin-Huxley equations. We found that the negative transient corresponded to an inward flow of sodium and the positive response to an outward flow of potassium. The field-effect transistors are able to probe the local flow of ionic currents in a membrane which is hidden in the region of cell adhesion. They may become a novel tool in neuroscience.

Animals↗
PubMed1997

Cable properties of dendrites in hippocampal neurons of the rat mapped by a voltage-sensitive dye.

Dendrites of pyramidal neurons from embryonic rat hippocampus are investigated in culture using a voltage-sensitive fluorescent dye. The electrical response to somatic stimulation is observed as a time-resolved map with a resolution of 0.9 microm at a time constant of 0.4 ms without signal averaging. The data are interpreted in terms of a tapering cable with Hodgkin-Huxley parametrization. The spread of short hyperpolarizing transients is damped by capacitive shunting. The invasion of an action potential is boosted by voltage-gated conductances of a low density. No irregularity is observed at a bifurcation. The passive cable parameters of internal resistance and membrane resistance at resting voltage are Ri = 300 omega cm and Rm = 40 (k)omega cm2 respectively, at a maximum sodium conductance of approximately 4.4 mS/cm2. The electrotonic length constant and the dynamic length constant at 1 kHz are 580 and 90 microm respectively. These results are compatible with electrophysiological data of dendrites in slices of adult hippocampus and with optical data of narrow processes of leech neurons in culture. The functional implications of boosting an action potential by voltage-gated channels of low density are considered.

Action Potentials↗
PubMed1994

Defined neuronal arborizations by guided outgrowth of leech neurons in culture.

Identified neurons of Hirudo medicinalis were cultivated on a protein extract of the extracellular matrix (ECM) of the leech. Microscopic patterns of active ECM protein were prepared by UV photolithography using aluminium masks. The shape of the patterns was visualized by a colour pattern formed in a dye-polymer substrate. The neurons were explanted on the root of branched ECM patterns. The patterns guided the outgrowth of neurites along linear lanes and they induced a bifurcation of the neurites under certain conditions. Neurons with a reproducible, regular shape of arborization were obtained within 1-2 days.

Animals↗
PubMed1994

Voltage-sensitive fluorescence of amphiphilic hemicyanine dyes in a black lipid membrane of glycerol monooleate.

Amphiphilic fluorescent hemicyanine dyes were adsorbed to a hemispherical bimolecular membrane of glycerol monooleate. Their excitation spectra of fluorescence were as in water, their emission spectra were as in hydrocarbon. An AC-voltage was applied across the membrane and the relative changes of the spectra of excitation and of emission were recorded. For all dyes we observed a blue-shift of excitation with positive voltage on the opposite side of staining. The effect is compared with the blue-shift expected for electrochromism. For most dyes we observed a red-shift of emission and a drop of the fluorescence intensity. These effects are compared with the red-shift and the drop of quantum yield expected for a voltage-induced solvatochromism caused by a minute displacement of the dyes in the anisotropic environment at the membrane/water interface.

Fluorescent Dyes↗
PubMed1994

Cable properties of a straight neurite of a leech neuron probed by a voltage-sensitive dye.

We measured a time-resolved map of electrical activity in a thin straight neurite (1.5 microns thick, 500 microns long) at a resolution of 8 microns and 0.4 ms. The neurite was obtained by guided outgrowth of an identified neuron of the leech on lanes of extracellular matrix protein. The electrical signals were detected by a fluorescent voltage-sensitive dye. We observed the voltage that was caused by an action potential elicited at the soma and by a Gaussian hyperpolarization induced at the soma, respectively. We compared the data with numerical solutions of the cable equation using the Hodgkin-Huxley parametrization. We could attribute the experimental results of depolarization and of hyperpolarization to the propagation of an action potential along an "active" cable and to the spread along a "passive" cable, respectively, if we assigned rather high specific resistances to the cytoplasm (RI = 250 omega.cm) and to the membrane (RM = 22 k omega.cm2). This assignment explained the slow velocity of 150 microns/ms of a pulse by active propagation and the limited range of 200 microns of a pulse by passive spread.

Action Potentials↗
PubMed1993

Voltage-sensitive fluorescence of amphiphilic hemicyanine dyes in neuron membrane.

Fluorescent amphiphilic hemicyanine dyes were adsorbed to the plasma membrane of isolated Retzius neurons of the leech. Voltage steps were applied to the neuron by the patch-clamp technique in whole-cell configuration. The change of fluorescence was observed as induced by the voltage jump. The relative changes of the excitation spectrum and of the emission spectrum of fluorescence were recorded. The complete set of spectral data for each dye was fitted by five parameters: shifts of the emission and the excitation spectrum, a change of fluorescence quantum yield and changes of the widths of the excitation and of the emission spectrum. The only common feature for all dyes was a blue-shift of the excitation spectrum and a drop of the yield when the neuron was stained from the outside and a positive voltage was applied to the inside. With respect to the shift of the emission spectrum and the changes of width qualitatively different results were obtained for different dyes. It is not attempted to assign a physical mechanism--probably a superposition of several mechanisms--of voltage-sensitivity.

Animals↗
PubMed1993

Exclusive-OR function of single arborized neuron.

We designed four arborized neurons which are able to evaluate the exclusive-or (XOR) function from two inputs. The input neurons form exclusively excitatory synapses on a dendritic tree which is a patchwork of "passive" (ohmic) and "active" cable segments. The active segments are described by the Hodgkin-Huxley model. The dynamics of the neurons and their output are obtained by numerical integration of the cable equation. In neurons 1 and 2 the XOR function is based on the annihilation of colliding action potentials. In neuron No. 3 the design takes advantage of the refractory period of action potentials. In neuron No. 4 voltage inversion is used as it occurs for inactivated sodium conductance in the Hodgkin-Huxley model. In all cases the XOR function depends critically on an appropriate timing of the input signals and on delays of the voltage transients in different branches of the dendrite.

Action Potentials↗
PubMed1992

Cable properties of arborized Retzius cells of the leech in culture as probed by a voltage-sensitive dye.

Retzius cells of Hirudo medicinalis were cultivated on extracellular matrix protein so that extended arborizations were formed. The propagation of voltage transients along 1-microns-thick neurites was observed at a resolution of 8 microns at 10 kHz by use of a voltage-sensitive dye. Delay and width of the fluorescence transients caused by hyperpolarization of the soma are described by passive spread of voltage in a homogeneous cable (time constant, 10 ms; space constant, 320 microns). The local sensitivity of the dye was determined from a comparison of the amplitudes of fluorescence and of fitted voltage. The fluorescence transients caused by depolarization were scaled using the sensitivity profile. Action potentials were found to pervade the neurites without significant change of amplitude but with enhanced pulse width.

Action Potentials↗
PubMed1991

Spectra of voltage-sensitive fluorescence of styryl-dye in neuron membrane.

The voltage sensitivity of fluorescence of an aminobenzstyryl-pyridinium dye (di4-ANEPPS) is characterized in Retzius cells dissociated from the leech. The modulation of the complete spectra of excitation and emission is determined. The spectral changes induced by depolarization are described by a blue shift of the absorption spectrum, by a weaker blue shift and an enhanced width of the fluorescence spectrum and by a decrease of the yield of fluorescence. These changes are attributed tentatively to a superposition of electrochromism and of field-induced resolvation.

Animals↗
PubMed1991

Guided outgrowth of leech neurons in culture.

Sensory neurons of Hirudo medicinalis were cultivated on patterns of extracellular matrix (ECM) protein which were prepared by UV irradiation using copper grid masks. The relation of the patterns and of the outgrowth of neurites was observed by scanning electronmicroscopy after gold decoration. The neurites were guided by narrow (10 microM) lanes of ECM protein. Branching of neurites was induced by branched lanes. Bent neurites were pulled off bent lanes at some distance behind the growth cone such that their length was reduced. Regularly spaced sites of adhesion became visible which remained connected to the neurites by extended filaments.

Animals↗