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G D Lange

Publications and source records attributed to G D Lange.

At least 19 recordsLinked to original sources
PubMed1994

Reciprocal expression of cell-cell coupling and voltage-dependent Na current during embryogenesis of rat telencephalon.

Using whole-cell patch-clamp techniques in situ (whole-tissue and tissue slices), we have studied two aspects of rat telencephalic cell development during the period of embryogenesis starting at E12. The first aspect was related to junctional coupling as revealed by low input resistance, intercellular dye spread and pharmacologic blockade. Coupling appeared to decrease with time, both in extent and occurrence. The second aspect dealt with cell excitability as revealed by voltage-dependent Na current (INa) expression. Immature action potentials and their underlying INaS were present in a small proportion of E12 cells. These currents were blocked 36% and 78% by 10(-7) M and 10(-6) M tetrodotoxin (TTX), respectively. From then onward, INaS got larger and more prevalent while no obvious changes in kinetics were observed. At E21, INaS were abolished by 10(-7) M TTX and channel density apparently was sufficient to support overshooting yet still immature action potentials.

Animals↗
PubMed1994

What does changing the temperature do to the melatonin rhythm in cultured chick pineal cells?

Chick pineal cells in static culture display a persistent, photosensitive circadian rhythm of melatonin production and release. We previously described the effects of light, the major physiological regulator of circadian rhythms, on the amplitude, period, and phase of the melatonin rhythm. Here we describe the effects of temperature, another physiological regulator of circadian rhythms, on the amplitude, period, and phase of this rhythm. Maintaining cells at 40.0-43.3 degrees C (104-110 degrees F) instead of 36.7 degrees C (98 degrees F) doubled the amplitude of the melatonin rhythm. In contrast, amplitude was reduced by about half at 33.3 degrees C (92 degrees F), and at 46.7 degrees C (116 degrees F) melatonin production was stopped within a few hours. Although temperatures of 40.0-43.3 degrees C raised melatonin output (unlike light, which suppresses it), they lengthened the period of the rhythm (as does constant light). Exposure of cells to 8-h pulses of these temperatures (40.0-43.3 degrees C) induced both phase delays and phase advances of the rhythm in subsequent cycles, with a phase dependence similar to that for the phase shifts induced by light pulses. Pulses of 40.0-43.3 degrees C were, however, weaker in their phase-shifting effects than light pulses. Pulses at still higher temperatures (46.7 degrees C) markedly inhibited melatonin output and delayed or disrupted the rhythm. The relationships (physiological and mechanistic) between the effects of temperature and light on the melatonin rhythm remain to be determined.

Animals↗
PubMed1993

Cl- channels are randomly activated by continuous GABA secretion in cultured embryonic rat hippocampal neurons.

Throughout the adult vertebrate central nervous system (CNS) gamma-aminobutyric acid (GABA) mediates transient Cl- conductances commonly identified as fast, Cl(-)-dependent inhibitory synaptic signals [Prog. Neurobiol., 36 (1991) 35-92]. In the rat hippocampus Cl(-)-dependent excitatory transients mediated by GABA emerge during the first postnatal week superimposed on a steady-state baseline that is also Cl(-)- and GABA-dependent [Int. J. Dev. Neurosci., 8 (1990) 481-490]. Here we report that many embryonic rat hippocampal neurons cultured for hours to days exhibit random fluctuations in Cl- channel activity that are mediated by continuous secretion of GABA in the absence of transients. Thus, GABA is broadcast tonically before it is released transiently.

Action Potentials↗
PubMed1993

GABAergic cells and signals appear together in the early post-mitotic period of telencephalic and striatal development.

Single cell suspensions derived from embryonic telencephala taken from embryos of gestational day 13 (E13) as well as rat striatal tissue from E14, 15 and 17 were prepared by tissue digestion with papain. Cell suspensions were analyzed by flow cytometry or plated onto poly-D-lysine-coated culture dishes for either nuclear staining or immunocytochemistry. Experiments on functional Na+ channels and GABAA receptor expression were carried out using a fluorescence-activated cell sorter (FACS) and a negatively charged fluorescent indicator dye (oxonol). FACS analysis of embryonic cell suspensions at E13-17 consistently revealed one major subpopulation accounting for 85-90% of the events and one minor subpopulation (10-15% of the total). When sorted, the major subpopulation consisted of phase-bright cells of 5-7 microns diameter some of which had neurites. The minor population consisted of phase-dark cells and resealed membranes of 0.5-4 microns diameter as well as debris. Almost all the cells obtained in the high FALS (forward-angle light scatter) subpopulation at E17 expressed 200-kDa neurofilament and tetanus toxin antigens while the small diameter cells seldom expressed tetanus toxin and particles never did. A small number of GABA-containing neurons were detected in the telencephalon at E13 (3%) and in the developing striatum at E14 (6%). All of the GABA-containing neurons expressed neurofilament. In the embryonic rat striatum, nanomolar concentrations of muscimol (GABAA agonist) induced depolarizing responses. A small number of cells in the high FALS subpopulation were responsive to muscimol starting at embryonic day 14, and the number of responsive cells increased at E15.(ABSTRACT TRUNCATED AT 250 WORDS)

Acridine Orange↗
PubMed1993

Ampullary sense organs, peripheral, central and behavioral electroreception in chimeras (Hydrolagus, Holocephali, Chondrichthyes).

Ampullary sense organs are distributed in groups over the head of Hydrolagus colliei with their pores in clusters and innervated by the buccal, hyomandibular and superficial ophthalmic branches of the anterior lateral line nerve. The ampullae contain ciliated sense cells in an alveolate-shaped epithelium, which communicates to the surface through a jelly-filled tube. The sense cells synapse at their bases with the afferent nerve fibers that terminate in the dorsal nucleus of the anterior lateral line lobe of the medulla. The anatomy and ultrastructure support the homology with the ampullae of Lorenzini of elasmobranchs. Single units recorded from the buccal branch of the anterior lateral line nerve are either lateral line or ampullary in character, the former being sensitive only to mechanical stimuli, the latter to both mechanical and to weak electric stimuli. They are also distinguished by the positions of their receptive fields. The electroreceptive units are spontaneously active and are excited by a cathode placed near the opening of their pore and inhibited by an anode. Compound evoked potentials are recorded from beneath the lateral aspect of the tectum in response to weak electric fields in the bath. Each recording locus has a best position and orientation of the electric field. The electric fields are effective if their duration is longer than ca. 2 ms; longer than 10 ms makes no difference until an OFF effect becomes distinct at ca. 50 ms. The reception is tuned to low frequencies but is not sensitive to maintained current (DC). Evoked potentials summating moderate numbers of responses are clear at < 1 microV/cm. Ratfish were conditioned in a ring-shaped tank to reverse the direction of swimming when an electric field was switched ON. The stimulus was a 5 Hz square wave or the onset of a DC of 1-10 microA between a pair of electrodes on the floor of the tank. The fish responded to fields as weak as 0.2 microV/cm. A specialized sense modality for electroreception, similar to that in elasmobranchs and most other groups of nonteleost fishes, except for Myxini and Neopterygii (holosteans), is present in the subclass Holocephali. The notion is supported that this modality and its central as well as peripheral apparatus arose early in the evolution of vertebrates. Only two losses of the whole system need be hypothesized, on this idea, once in the ancestors of the hagfishes and once in the ancestors of the neopterygians, which include the teleosts. Some orders of teleosts then evolved a new system of electroreception independently. The ciliary receptor cells are probably primitive; microvillar sense cells evolved independently.

Animals↗
PubMed1993

Characterization of the gamma-aminobutyric acidA receptor-channel complex composed of alpha 1 beta 2 and alpha 1 beta 3 subunits from rat brain.

The cloned alpha 1, beta 2 and beta 3 subunits of the gamma-aminobutyric acid (GABA)A receptor-channel complex from rat brain were coexpressed as alpha beta complexes in cultured Chinese hamster ovary cells. Electrophysiological characterization of alpha 1 beta 2 and alpha 1 beta 3 receptor subunit arrangements was performed utilizing patch electrodes in the whole-cell recording configuration. The reversal potential of the current activated by either GABA or muscimol corresponded to that expected for Cl- ions and was dependent on the Cl- gradient. The dose response to GABA for activation of Cl- currents by either subunit combination displayed similar potencies. Currents were partially blocked by the reversible antagonist bicuculline. (-)Pentobarbital was ineffective by itself, but potentiated responses to GABA. The steroid alphaxalone (3 alpha-hydroxy 5 alpha-pregnane 11,20-dione) produced just-detectable inward currents, but did not potentiate GABA-activated currents. Diazepam was completely ineffective. The kinetics and conductance of the Cl- ion channels were inferred from spectral analysis of agonist-induced current fluctuations. Both kinetics and conductance were dependent on agonist structure.

Animals↗
PubMed1991

A fractal analysis of pyramidal neurons in mammalian motor cortex.

Pyramidal neurons in the mammalian cerebral cortex can be described by a fractal dimension (Mandelbrot, 1982), which is an objective, quantitative measure of the complexity of their soma/dendritic borders. In the cat, the fractal dimensions of lamina V cells, which include pyramidal tract neurons (PTN), indicate that these cells are more complex than other pyramidal neurons (PN) in the same region of motor cortex. The lamina V cells of the cat are also more complex than those in motor cortex of the monkey. Moreover, lamina III neurons in the monkey are more complex than monkey lamina V neurons. The fractal dimension of the intracortical axon collateral arborizations of the same pyramidal neurons indicated, in all cases, that the branching of these terminals is less complex than the branching of the dendrites of the same cells. In line with the observation that the fractal dimensions of some homologous cellular populations are different in different species, it is suggested that the fractal dimension and the degree of morphological complexity may relate to the requirement for the number of separable functions to be accommodated within one neuron. For example, as the size of the cortex and the number of neurons in a region increase, the opportunity exists within a given cortical zone, for individual functions to be segregated and for functional specialization to be accommodated with less morphological complexity of the individual neurons performing each of these functions.

Animals↗
PubMed1991

A fractal analysis of cultured rat optic nerve glial growth and differentiation.

Fractal dimension can be used as a quantitative measure of morphological complexity. Separate, enriched populations of oligodendrocytes or type 2 astrocytes derived from neonatal rat optic nerves were allowed to differentiate in vitro. Fractal dimensions of differentiating glial cells were measured over time. The fractal dimension correlated with perceived complexity and increased in value as the glial cells matured. Analysis of the changes in fractal dimension with time revealed unique rates of growth and differentiation for each glial phenotype.

Animals↗
PubMed1990

Electrical and chemical excitability appear one week before birth in the embryonic rat spinal cord.

Embryonic rat spinal cord cells were acutely dissociated with the enzyme papain, stained with a voltage-sensitive oxonol dye and incubated with various pharmacological agents. Changes in the fluorescence intensity and, by inference, membrane potential of the cells were analyzed in a flow cytometer. Veratridine caused depolarization of the cells in a TTX-sensitive manner from as early as embryonic day 13. Depolarizing responses to muscimol and kainate appeared slightly later, at embryonic days 14 and 15, and were blocked by the antagonists bicuculline and CNQX, respectively. Responses to veratridine and kainate did not occur in sodium-free medium. The emergence of these excitable membrane properties coincides with postmitotic differentiation and synaptic development in the embryonic spinal cord.

Animals↗
PubMed1990

Embryonic and early postnatal hippocampal cells respond to nanomolar concentrations of muscimol.

Embryonic and early postnatal tissue taken from rat hippocampi were papain digested in order to obtain cell suspensions suitable for analysis in a fluorescence-activated cell sorter (FACS). Cell suspensions consisted of two major peaks of forward-angle light scatter (FALS). FACS analysis showed that the population which stained intensely with the vital dye Acridine orange (AO) scattered significant levels of light (high FALS) and amounted to 85% of the total events collected in embryonic cell suspensions and 65% in postnatal (PN) samples. Two minor populations were weakly stained with AO and scattered little light. Oxonol, a voltage-sensitive indicator dye, was used to detect membrane polarization changes. The AO and oxonol staining patterns were very similar. All the events exposed to media containing 50 mM KCl were depolarized (increase in intensity of oxonol fluorescence). The depolarizing effect of veratridine, a sodium channel activator, was more pronounced in the high FALS subpopulation. In embryonic hippocampal cell suspensions nanomolar concentrations of GABAA agonists depolarized the high FALS subpopulation in a dose-dependent manner. This effect was prevented by preincubation with bicuculline or picrotoxin. In hippocampal cell suspensions obtained from 5-7-day-old rat pups (PN5-7), GABAA agonists depolarized one cell subpopulation and hyperpolarized another. Our results indicate that physiological responses can be resolved in subpopulations of hippocampal cell suspensions by FACS analysis. This technique seems to be a sensitive assay to measure physiological responses (changes in membrane potential) as a parameter of receptor expression. GABAA agonists induced pure depolarizing responses in embryonic and early postnatal hippocampus when active neurogenesis is taking place. The response become hyperpolarizing-depolarizing ones after inhibitory synapses appear.

Aging↗
PubMed1989

A fractal analysis of cell images.

Methods of digital image analysis have been adapted to measure the fractal dimension of cellular profiles. The fractal dimension is suggested as a useful measure of the complexity of a contour. Three methods produce similar results when applied to constructed, near-ideal fractal figures. Comparison of the measurements for a variety of image types indicates the measurement accuracy in each case and may help in interpreting the results when applied to real, non-ideal cell images of unknown fractal dimension. Two of the methods are currently adopted as appropriate for use on neuronal contours. A correlation exists between the complexity of these contours and the magnitude of the estimated fractal dimension.

Animals↗
PubMed1988

(-)-Baclofen activates presynaptic GABAB receptors on GABAergic inhibitory neurons from embryonic rat hippocampus.

GABA-mediated monosynaptic inhibitory postsynaptic currents were recorded at synapses between embryonic rat hippocampal neurons in culture. The GABAB receptor agonist L(-)-baclofen reduced inhibitory postsynaptic current amplitude substantially at 10 microM, while the D(+)-isomer of baclofen was without effect. L(-)-Baclofen did not increase postsynaptic membrane conductance or block postsynaptic membrane responses to GABA. It is concluded that L(-)-baclofen depresses GABA-mediated inhibitory postsynaptic potentials by a presynaptic action at GABAB receptors on or near the terminals of the inhibitory neuron. The possible physiological importance of these presynaptic 'autoreceptors' is discussed.

Action Potentials↗
PubMed1988

Flow cytometric analysis of membrane potential in embryonic rat spinal cord cells.

Flow cytometric analysis of membrane potential in suspensions of embryonic rat spinal cord cells was carried out in a fluorescence-activated cell sorter (FACS) using anionic voltage-sensitive, fluorescent dyes (oxonols). The FACS or flow cytometer is an analytical instrument that measures optical properties of large cell populations at a single cell level of resolution. The incorporation of oxonol allows relative measurements of membrane potential, since the partition of oxonol within the plasmalemma is directly related to the degree of cell depolarization. Incubation of cells in elevated K+ concentrations or with the Na+ channel agonist batrachotoxin (BTX) changed the fluorescence intensity distribution pattern of the live-cell population; these changes were consistent with the depolarizing effects of these manipulations. Fluorescence shifts were either undetectable or minimal in the dead-cell population. The BTX-induced shift was blocked by tetrodotoxin (TTX) and was reversed in Na+-free medium, indicating embryonic expression of functional Na+ channels. Fluorescence microscopy of sorted cells showed that live cells typically exhibited circumferential ring-like patterns, whose intensities were enhanced under depolarizing conditions. The results show that flow cytometry combined with oxonol dyes can be used to measure the relative membrane potential of large numbers of individual central nervous system cells. The analysis of the changes in the distributions of these membrane potentials can be used to reveal the development of functional ion conductance mechanisms.

Animals↗
PubMed1988

Edge detection in images using Marr-Hildreth filtering techniques.

Details of the morphology of light microscope images of horseradish peroxidase labeled mammalian neurons in cell culture were investigated. A modified Marr-Hildreth edge-detecting algorithm was used in an image processor to obtain a continuous border of the labeled neurons. The interior of the border was filled to obtain isolated binary silhouettes of the neurons. These silhouettes can be used for further quantitative studies.

Animals↗
PubMed1987

Potentiation of gamma-aminobutyric-acid-activated chloride conductance by a steroid anaesthetic in cultured rat spinal neurones.

1. Intracellular recordings from cultured rat spinal cord neurones demonstrated that Cl(-)-dependent responses to GABA (gamma-aminobutyric acid) (but not glycine) were increased in amplitude and duration by the steroid anaesthetic alphaxalone (3 alpha-hydroxy-5 alpha-pregnane-11,20-dione) at submicromolar concentrations that produced little or no effect on passive electrical properties. The non-anaesthetic 3 beta-hydroxy analogue was without effect on GABA-evoked responses. 2. Under voltage clamp, membrane currents evoked by GABA were potentiated by alphaxalone without change in the reversal potential for the GABA-evoked response. Fluctuation analysis of GABA-evoked currents suggested that the mean open-time of GABA-activated channels was prolonged from 30 to 74 ms in the presence of the anaesthetic. 3. Higher concentrations of alphaxalone, similar to those reported during surgical anaesthesia, increased membrane conductance in the absence of exogenously applied GABA. Under voltage clamp, current responses to alphaxalone reversed at the same potential as did responses to GABA, suggesting that they result from increased Cl- conductance. 4. Alphaxalone responses were reduced by the GABA antagonist bicuculline. Fluctuation analysis of current responses to the anaesthetic suggest that they result from the activation of ion channels of long (100 ms) open-time and elementary conductance indistinguishable from that of channels activated by GABA (20 pS). Taken together, these findings indicate that the steroid anaesthetic is able to directly activate Cl- conductance normally activated by GABA in spinal neurones. 5. The actions of the steroid at GABA-receptor-Cl(-)-channel complexes are similar to those produced by the anaesthetic barbiturates (e.g. pentobarbitone), although obtained at 50-100-fold lower concentrations. These effects on the inhibitory Cl(-)-conductance mechanism may be partly responsible for the depressant actions of alphaxalone on the mammalian central nervous system.

Anesthetics↗
PubMed1987

The influence of skeletal muscle on the electrical excitability of dorsal root ganglion neurons in culture.

Dorsal root ganglion (DRG) neurons from embryonic mice grown in coculture with dissociated skeletal muscle or in skeletal muscle conditioned medium (CM) showed an increased incidence of repetitive firing of action potentials when injected with sustained (60-100 msec) depolarizing current. This is in contrast to DRG neurons grown in monoculture and normal medium, which exhibit such behavior far less frequently. The first action potential showed less sensitivity to block with TTX and more sensitivity to Ca2+ channel blockers than the subsequent action potentials. The increased incidence of repetitive firing occurred when CM was added after as few as 2 or as many as 22 d in culture and with as little as 1-7 hr exposure to CM. This effect of CM cannot be mimicked by NGF or by coculture with cells from embryonic spinal cord (Peacock et al., 1973), can be eliminated by heating the CM at 56 degrees C for 30 min, and partially reversed following short exposure to CM. These results indicate that skeletal muscle releases some heat-labile factor(s) that can cause repetitive firing and, in addition, significant decrease in input resistance in the CM-treated neurons and a depression of the anomalous rectification, neither of which could account for the increase in repetitive firing.

Action Potentials↗
PubMed1986

Analysis and isolation of embryonic mammalian neurons by fluorescence-activated cell sorting.

Cells were dissociated from the CNS of the embryonic mouse and rat to produce cell suspensions suitable for analysis and separation on a fluorescence-activated cell sorter (FACS). Cells from the spinal cord of the embryonic mouse were analyzed in the most detail. Cell suspensions generated three major peaks in histograms of forward-angle light scatter. Examination of material isolated from each peak and labeling of cell suspensions with the nonvital and supravital fluorescent dyes propidium iodide, ethidium bromide, and acridine orange demonstrated that the three peaks represented live cells, dead cells, and subcellular fragments. Passage through the cell sorter did not detectably damage live cells, as shown by light microscopy, FACS analysis, and in vitro culture of sorted cells. Neurons and glial cells collected by sorting survived at least 4 weeks in culture. Cell suspensions dissociated from the dorsal root ganglia, hippocampus, hypothalamus, cerebellum, and cerebral cortex of the embryonic mouse and from the spinal cord of the embryonic rat produced similar results. Analysis of samples prepared at different developmental stages showed that viable cells could be recovered from each of these regions throughout the important stages of neurogenesis and early cellular differentiation, but that few viable cells could be recovered from animals beyond late embryonic or early postnatal ages. Quantitative FACS analysis of monoclonal antibody A2B5, tetanus toxin and cholera toxin, and lectins binding to live dissociated cells from the embryonic spinal cord demonstrated that these cells had already developed binding sites for these cell-surface ligands by embryonic day 13. These results demonstrate that a fluorescence-activated cell sorter can be used for quantitative analysis of specific cellular properties, that FACS analysis and sorting can be used to identify and isolate live cells from many regions of the embryonic mammalian CNS during important developmental periods, and that sorted neurons and glial cells can be maintained for weeks in culture.

Animals↗