Pharmacological Analysis of Tonic Activity in Motoneurons During Stick Insect Walking

2009 ◽  
Vol 102 (2) ◽  
pp. 1049-1061 ◽  
Author(s):  
Sandra Westmark ◽  
Eugenio E. Oliveira ◽  
Joachim Schmidt
Keyword(s):  
Second Messengers ◽  
Cell Voltage ◽  
Stick Insect ◽  
Excitatory Input ◽  
Inward Current ◽  
Camp Analogue ◽  
Bath Application ◽  
Premotor Neurons

Stick insect middle leg (mesothoracic) motoneurons receive tonic excitatory input during front leg stepping on a treadmill. We studied the pharmacology of this excitatory input to the motoneurons during single-legged treadmill walking (in situ). During bath application of drugs restricted to the mesothoracic ganglion, activity in motoneurons contralateral to the stepping front leg was recorded from neuropilar processes. Application of the cholinergic antagonist atropine reduced the tonic depolarization amplitude. These results were compared with findings in acutely dissociated motoneuron cell bodies (in vitro) under whole cell voltage-clamp conditions. The presence of an acetylcholine-induced current in situ was supported by the finding of an acetylcholine evoked biphasic inward current with a sustained component that could be blocked by atropine. In situ the tonic depolarization was generally increased by application of the neuro-modulator octopamine and decreased by its antagonist mianserin. In vitro, however, octopamine reduced the inward current evoked by acetylcholine application to motoneurons. Intracellular application of bis-( o-aminophenoxy)- N,N,N',N'-tetraacetic acid (BAPTA) into motoneurons in situ revealed a dependence of the tonic depolarization on Ca2+ and application of the membrane-permeable cAMP analogue 8-bromo-cAMP increased the tonic depolarization. In contrast, 8-bromo-cAMP reduced the inward current evoked by acetylcholine application to motoneurons in vitro. We conclude that during walking, acetylcholine contributes to mediating the tonic depolarization possibly by acting on atropine-sensitive receptors on motoneurons. Octopamine that is released during walking increases the tonic depolarization. This increase, however, is not based on modulation of cholinergic action on motoneurons but rather on effects on premotor neurons. Both, Ca2+ and cAMP are likely second messengers involved in mediating the tonic depolarization, but whereas Ca2+ acts in motoneurons, cAMP does not appear to mediate a cholinergic depolarization in motoneurons.

scholarly journals Noradrenergic and GABAB Receptor Activation Differentially Modulate Inputs to the Premotor Nucleus RA in Zebra Finches

2008 ◽  
Vol 100 (1) ◽  
pp. 8-18 ◽  
Author(s):  
Max Sizemore ◽  
David J. Perkel
Keyword(s):  
Gabab Receptor ◽  
Cell Voltage ◽  
Receptor Activation ◽  
Social Contexts ◽  
Excitatory Input ◽  
Song Learning ◽  
Zebra Finches ◽  
Proper Name ◽  
Magnocellular Nucleus

Neuromodulators can rapidly modify neural circuits, altering behavior. Songbirds provide an excellent system for studying the role of neuromodulation in modifying circuits that underlie behavior because song learning and production are mediated by a discrete set of interconnected nuclei. We examined the neuromodulatory effects of noradrenergic and GABAB receptor activation on synaptic inputs to the premotor robust nucleus of the arcopallium (RA) in zebra finches using whole cell voltage-clamp recording in vitro. In adults, norepinephrine strongly reduced input from the lateral magnocellular nucleus of the anterior nidopallium (LMAN) but only slightly reduced the input from nucleus HVC (proper name), the excitatory input from axon collaterals of other RA neurons, and input from GABAergic interneurons. The effect of norepinephrine was mimicked by the α2 adrenoceptor agonist UK14,304 and blocked by the α2 antagonist yohimbine. Conversely, the GABAB receptor agonist baclofen strongly decreased HVC, collateral, and GABAergic inputs to RA neurons while causing little reduction in the LMAN input. In juveniles undergoing song learning, norepinephrine reduced the LMAN input, caused only a small reduction in the HVC input, and greatly reduced the collateral and GABAergic inputs. Baclofen caused similar results in juvenile and adult birds, reducing HVC, collateral, and GABAergic inputs significantly more than the LMAN input. Significant increases in paired-pulse ratio accompanied all reductions in synaptic transmission, suggesting a presynaptic locus. The reduction in the LMAN input by norepinephrine may be important for mediating changes in song elicited by different social contexts and is well-placed to play a role in song learning.

Neurotoxicity Mediated by Aberrant Patterns of Synaptic Activity Between Rat Hippocampal Neurons in Culture

1998 ◽  
Vol 80 (5) ◽  
pp. 2688-2698 ◽  
Author(s):  
John R. McLeod ◽  
Maoxing Shen ◽  
Daniel J. Kim ◽  
Stanley A. Thayer
Keyword(s):  
Cell Death ◽  
Nmda Receptor ◽  
Receptor Antagonist ◽  
Hippocampal Neurons ◽  
Cell Voltage ◽  
Nmda Receptor Antagonist ◽  
Synaptic Activity ◽  
Slow Inward Current ◽  
Inward Current

McLeod, John R., Jr., Maoxing Shen, Daniel J. Kim, and Stanley A. Thayer. Neurotoxicity mediated by aberrant patterns of synaptic activity between rat hippocampal neurons in culture. J. Neurophysiol. 80: 2688–2698, 1998. Reducing the extracellular Mg2+ concentration ([Mg2+]o) to 0.1 mM evoked an aberrant pattern of glutamatergic activity in the synaptic network formed by rat hippocampal neurons grown in primary culture. This treatment resulted in a significant increase in neuronal death when maintained for 20–24 h; 0.1 mM [Mg2+]o elicited a stable and repetitive series of intracellular Ca2+ concentration ([Ca2+]i) spikes as indicated by indo-1-based microfluorimetry. Fura-2-based digital imaging experiments found that the [Ca2+]i spikes were synchronized for all the neurons in a given field. Thus electrophysiological recordings from individual cells were reasonable representations of the field as a whole, enabling correlation of electrical activity to viability. Underlying each [Ca2+]i spike was an intense burst of action potentials. Whole cell voltage-clamp experiments showed that a burst was composed of fast action currents superimposed on a slow inward current. The N-methyl-d-aspartate (NMDA) receptor antagonist CGS19755 (10 μM) blocked [Ca2+]i spiking, the slow inward current, and the cell death induced by low [Mg2+]o. The L-type Ca2+ channel antagonist nimodipine (10 μM) blocked [Ca2+]i spiking, all synaptic activity, and the cell death induced by low [Mg2+]o. The non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 μM) exerted variable effects on [Ca2+]i spiking and blocked the slow inward current only when the cells were held at a relatively negative holding potential. CNQX did not afford any protection from 0.1 mM [Mg2+]o-induced neurotoxicity. [Ca2+]i imaging experiments showed that CNQX inhibited [Ca2+]i spiking in a subset of neurons within an active network. Thus, the neurons that were insensitive to CNQX appear to be those that were destined to die. We characterized an in vitro model that allowed us to correlate specific electrophysiological components of glutamatergic synaptic activity to the subsequent viability of the network. A slow NMDA receptor-mediated inward current was required to elicit [Ca2+]i spiking and neurotoxicity. Non-NMDA receptors did not contribute to synaptically mediated cell death in this model. An L-type Ca2+ channel antagonist was neuroprotective when used at concentrations that blocked synaptic activity, suggesting that dendritic L-type Ca2+ channels present a useful target for neuroprotective drugs.

Palmitoyl carnitine modifies sodium currents and induces transient inward current in ventricular myocytes

1994 ◽  
Vol 266 (3) ◽  
pp. H1034-H1046 ◽  
Author(s):  
J. Wu ◽  
P. B. Corr
Keyword(s):  
Ventricular Myocytes ◽  
Cell Voltage ◽  
Ionic Currents ◽  
Inward Current ◽  
Palmitoyl Carnitine ◽  
Transient Inward Current ◽  
Ventricular Cells ◽  
Current Voltage

Long-chain acylcarnitines increase within 2 min in ischemic myocardium in vivo and induce delayed afterdepolarizations (DADs) and complex oscillations of membrane potential in vitro. This study was performed to assess the ionic currents underlying these electrophysiological alterations in isolated rabbit ventricular cells using whole cell voltage-clamp procedures. Palmitoyl carnitine (10 microM, for 6-10 min) elicited a transient inward current (Iti) in the presence of blockade of Ca2+ and K+ channels. The effect of palmitoyl carnitine was reversible after washout (n = 6). The amplitude of Iti was dependent on the amplitude of the preceding depolarization step. Palmitoyl carnitine (10 microM, for > 2 min) also induced another inward current, which was activated spontaneously at potentials between -120 and -20 mV with a linear current-voltage relationship (1.0 +/- 0.1 nA at -80 mV). This current was abolished by replacing extracellular Na+ with tetraethylammonium chloride, indicating that Na+ was the charge carrier. Inactivation of this current was slow (gamma = 885.9 +/- 89.1 ms, n = 12) or incomplete, indicating the appearance of a slow-inactivating Na+ inward current [INa(s)]. Palmitoyl carnitine always induced INa(s) before the appearance of Iti. Intracellular ethylene glycol-bis(beta-amino-ethyl ether)-N,N,N',N'-tetraacetic acid (10 mM) abolished Iti but did not suppress INa(s) (n = 4), indicating that INa(s) was not activated by intracellular Ca2+ (Cai2+). Tetrodotoxin (10 microM) also decreased the amplitude of INa(s). Thus palmitoyl carnitine induces INa(s), which likely leads to an increase in Na+ influx, thereby eliciting an increase in Cai2+ via the Na(+)-Ca2+ exchanger and leading to the development of Iti, DADs, and triggered activity.

In situ determination of [Ca2+]i threshold for translocation of the alpha-protein kinase C isoform

1994 ◽  
Vol 266 (6) ◽  
pp. C1544-C1551 ◽  
Author(s):  
R. A. Khalil ◽  
C. Lajoie ◽  
K. G. Morgan
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Second Messengers ◽  
In Vitro Assays ◽  
Intact Cells ◽  
Kinase C ◽  
Biochemical Assays

Because of inherent difficulties in maintaining physiological conditions in biochemical assays, the intracellular free Ca2+ concentration ([Ca2+]i) required for activation of protein kinase C (PKC) in intact cells remains unclear. In the present study, [Ca2+]i was measured in freshly isolated vascular smooth muscle cells loaded with fura 2 while, in parallel, the distribution of the Ca(2+)-dependent alpha-PKC isoform was monitored using digital imaging microscopy. The [Ca2+]i alpha-PKC translocation threshold was determined by changing extracellular free Ca2+ concentration in steps while monitoring [Ca2+]i. In the absence of agonists, increasing [Ca2+]i caused < 25% of maximal translocation. In the presence of phenylephrine, maximum translocation occurred at [Ca2+]i > or = 198 nM. Phenylephrine augmented translocation of alpha-PKC primarily by increasing the slope of the [Ca2+]i-PKC translocation relationship. These results indicate that the [Ca2+]i threshold of alpha-PKC translocation in situ is less than that reported in most in vitro assays and are consistent with an effect of agonist-induced generation of other second messengers that cause cooperative interactions leading to translocation.

Correlated Studies of Cellular Interactions Using TEM, Freeze Etching, and SEM

1974 ◽  
Vol 32 ◽  
pp. 320-321
Author(s):  
J. P. Revel
Keyword(s):  
Developmental Stages ◽  
Three Dimensional ◽  
Cell Movement ◽  
Cellular Interactions ◽  
Serial Sections ◽  
Cell Sheets ◽  
Freeze Etching ◽  
Early Developmental

Movement of individual cells or of cell sheets and complex patterns of folding play a prominent role in the early developmental stages of the embryo. Our understanding of these processes is based on three- dimensional reconstructions laboriously prepared from serial sections, and from autoradiographic and other studies. Many concepts have also evolved from extrapolation of investigations of cell movement carried out in vitro. The scanning electron microscope now allows us to examine some of these events in situ. It is possible to prepare dissections of embryos and even of tissues of adult animals which reveal existing relationships between various structures more readily than used to be possible vithout an SEM.

In vitro and in vivo polysaccharides assembly: ultrastructural and cytochemical study of growing plant cell wall components.

1978 ◽  
Vol 36 (2) ◽  
pp. 434-435
Author(s):  
D. Reis ◽  
B. Vian ◽  
J. C. Roland
Keyword(s):  
Cell Wall ◽  
Self Assembly ◽  
Plant Cell Wall ◽  
Higher Plants ◽  
Three Dimensional ◽  
Cytochemical Study ◽  
Wall Components

Wall morphogenesis in higher plants is a problem still open to controversy. Until now the possibility of a transmembrane control and the involvement of microtubules were mostly envisaged. Self-assembly processes have been observed in the case of walls of Chlamydomonas and bacteria. Spontaneous gelling interactions between xanthan and galactomannan from Ceratonia have been analyzed very recently. The present work provides indications that some processes of spontaneous aggregation could occur in higher plants during the formation and expansion of cell wall.Observations were performed on hypocotyl of mung bean (Phaseolus aureus) for which growth characteristics and wall composition have been previously defined.In situ, the walls of actively growing cells (primary walls) show an ordered three-dimensional organization (fig. 1). The wall is typically polylamellate with multifibrillar layers alternately transverse and longitudinal. Between these layers intermediate strata exist in which the orientation of microfibrils progressively rotates. Thus a progressive change in the morphogenetic activity occurs.

In Situ Localization of PPAR Gamma and Uncoupling Protein in Mouse Embryo Sections Using Digoxigenin-Labeled Riboprobes

1996 ◽  
Vol 54 ◽  
pp. 32-33
Author(s):  
C. Jennermann ◽  
S. A. Kliewer ◽  
D. C. Morris
Keyword(s):  
Alkaline Phosphatase ◽  
Room Temperature ◽  
Uncoupling Protein ◽  
Ppar Gamma ◽  
Nuclear Hormone Receptor ◽  
Full Length ◽  
Northern Analysis ◽  
Peroxisome Proliferator

Peroxisome proliferator-activated receptor gamma (PPARg) is a member of the nuclear hormone receptor superfamily and has been shown in vitro to regulate genes involved in lipid metabolism and adipocyte differentiation. By Northern analysis, we and other researchers have shown that expression of this receptor predominates in adipose tissue in adult mice, and appears first in whole-embryo mRNA at 13.5 days postconception. In situ hybridization was used to find out in which developing tissues PPARg is specifically expressed.Digoxigenin-labeled riboprobes were generated using the Genius™ 4 RNA Labeling Kit from Boehringer Mannheim. Full length PPAR gamma, obtained by PCR from mouse liver cDNA, was inserted into pBluescript SK and used as template for the transcription reaction. Probes of average size 200 base pairs were made by partial alkaline hydrolysis of the full length transcripts. The in situ hybridization assays were performed as described previously with some modifications. Frozen sections (10 μm thick) of day 18 mouse embryos were cut, fixed with 4% paraformaldehyde and acetylated with 0.25% acetic anhydride in 1.0M triethanolamine buffer. The sections were incubated for 2 hours at room temperature in pre-hybridization buffer, and were then hybridized with a probe concentration of 200μg per ml at 70° C, overnight in a humidified chamber. Following stringent washes in SSC buffers, the immunological detection steps were performed at room temperature. The alkaline phosphatase labeled, anti-digoxigenin antibody and detection buffers were purchased from Boehringer Mannheim. The sections were treated with a blocking buffer for one hour and incubated with antibody solution at a 1:5000 dilution for 2 hours, both at room temperature. Colored precipitate was formed by exposure to the alkaline phosphatase substrate nitrobluetetrazoliumchloride/ bromo-chloroindlylphosphate.

CONSERVATION OF RARE SPECIES PLANTS AND LICHENS IN SITU WITH PLANNED ANTHROPOGENIC ACTIVITIES: BASIC APPROACHES AND IMPLEMENTATION PRINCIPLES

2018 ◽  
Vol 12 (7-8) ◽  
pp. 38-45
Author(s):  
A. N. EFREMOV ◽  
N. V. PLIKINA ◽  
T. ABELI
Keyword(s):  
Rare Species ◽  
Technology Implementation ◽  
Technology Development ◽  
Anthropogenic Activities ◽  
Natural Habitat ◽  
Local Population ◽  
Main Stage ◽  
Social Risks

Rare species are most vulnerable to man-made impacts, due to their biological characteristics or natural resource management. As a rule, the economic impact is associated with the destruction and damage of individual organisms, the destruction or alienation of habitats. Unfortunately, the conservation of habitat integrity is an important protection strategy, which is not always achievable in the implementation of industrial and infrastructural projects. The aim of the publication is to summarize the experience in the field of protection of rare species in the natural habitat (in situ), to evaluate and analyze the possibility of using existing methods in design and survey activities. In this regard, the main methodological approaches to the protection of rare species in the natural habitat (in situ) during the proposed economic activity were reflected. The algorithm suggested by the authors for implementing the in situ project should include a preparatory stage (initial data collection, preliminary risk assessments, technology development, obtaining permitting documentation), the main stage, the content of which is determined by the selected technology and a long monitoring stage, which makes it possible to assess the effectiveness of the taken measures. Among the main risks of in situ technology implementation, the following can be noted: the limited resources of the population that do not allow for the implementation of the procedure without prior reproduction of individuals in situ (in vitro); limited knowledge of the biology of the species; the possibility of invasion; the possibility of crossing for closely related species that сo-exist in the same habitat; social risks and consequences, target species or population may be important for the local population; financial risks during the recovery of the population. The available experience makes it possible to consider the approach to the conservation of rare species in situ as the best available technology that contributes to reducing negative environmental risks.

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