ABSTRACT
Homeostatic mechanisms regulate synaptic function to maintain nerve and muscle excitation within reasonable physiological limits. The mechanisms that initiate homeostasic changes to synaptic function are not known. We specifically impaired cellular depolarization by expressing the Kir2.1 potassium channel in Drosophila muscle. In Kir2.1-expressing muscle there is a persistent outward potassium current ( approximately 10 nA), decreased muscle input resistance (50-fold), and a hyperpolarized resting potential. Despite impaired muscle excitability, synaptic depolarization of muscle achieves wild-type levels. A quantal analysis demonstrates that increased presynaptic release (quantal content), without a change in quantal size (mEPSC amplitude), compensates for altered muscle excitation. Because morphological synaptic growth is normal, we conclude that a homeostatic increase in presynaptic release compensates for impaired muscle excitability. These data demonstrate that a monitor of muscle membrane depolarization is sufficient to initiate synaptic homeostatic compensation.
Subject(s)
Excitatory Postsynaptic Potentials/physiology , Homeostasis/physiology , Potassium Channels, Inwardly Rectifying , Potassium Channels/metabolism , Presynaptic Terminals/metabolism , Synaptic Transmission/physiology , Animals , Drosophila , Gene Expression/physiology , Membrane Potentials/physiology , Motor Neurons/physiology , Muscles/innervation , Muscles/physiology , Potassium/metabolism , Potassium Channels/genetics , Receptors, Glutamate/metabolismABSTRACT
The precise regulation of neural excitability is essential for proper nerve cell, neural circuit, and nervous system function. During postembryonic development and throughout life, neurons are challenged with perturbations that can alter excitability, including changes in cell size, innervation, and synaptic input. Numerous experiments demonstrate that neurons are able to compensate for these types of perturbation and maintain appropriate levels of excitation. The mechanisms of compensation are diverse, including regulated changes to synaptic size, synaptic strength, and ion channel function in the plasma membrane. These data are evidence for homeostatic regulatory systems that control neural excitability. A model of neural homeostasis suggests that information about cell activity, cell size, and innervation is fed into a system of cellular monitors. Intracellular- and intercellular-signaling systems transduce this information into regulated changes in synaptic and ion channel function. This review discusses evidence for such a model of homeostatic regulation in the nervous system.
Subject(s)
Homeostasis/physiology , Neuronal Plasticity/physiology , Neurons/physiology , Synapses/physiology , AnimalsABSTRACT
We present evidence that Futsch, a novel protein with MAP1B homology, controls synaptic growth at the Drosophila neuromuscularjunction through the regulation of the synaptic microtubule cytoskeleton. Futsch colocalizes with microtubules and identifies cytoskeletal loops that traverse the lateral margin of select synaptic boutons. An apparent rearrangement of microtubule loop architecture occurs during bouton division, and a genetic analysis indicates that Futsch is necessary for this process. futsch mutations disrupt synaptic microtubule organization, reduce bouton number, and increase bouton size. These deficits can be partially rescued by neuronal overexpression of a futsch MAP1B homology domain. Finally, genetic manipulations that increase nerve-terminal branching correlate with increased synaptic microtubule loop formation, and both processes require normal Futsch function. These data suggest a common microtubule-based growth mechanism at the synapse and growth cone.
Subject(s)
Drosophila Proteins , Drosophila/physiology , Microtubule-Associated Proteins/physiology , Microtubules/physiology , Nerve Growth Factors/physiology , Synapses/physiology , Animals , Cytoskeleton/metabolism , Cytoskeleton/ultrastructure , Drosophila/growth & development , Drosophila/metabolism , Microtubules/ultrastructure , Nerve Endings/physiology , Nerve Endings/ultrastructure , Presynaptic Terminals/metabolism , Presynaptic Terminals/ultrastructure , Synapses/ultrastructure , Tissue DistributionABSTRACT
Postsynaptic dCBP (Drosophila homolog of the CREB binding protein) is required for presynaptic functional development. Viable, hypomorphic dCBP mutations have a approximately 50% reduction in presynaptic transmitter release without altering the Ca2+ cooperativity of release or synaptic ultrastructure (total bouton number is increased by 25%-30%). Exogenous expression of dCBP in muscle rescues impaired presynaptic release in the dCBP mutant background, while presynaptic dCBP expression does not. In addition, overexpression experiments indicate that elevated dCBP can also inhibit presynaptic functional development in a manner distinct from the effects of dCBP loss of function. Pre- or postsynaptic overexpression of dCBP (in wild type) reduces presynaptic release. However, we do not observe an increase in bouton number, and presynaptic overexpression impairs short-term facilitation. These data suggest that dCBP participates in a postsynaptic regulatory system that controls functional synaptic development.
Subject(s)
Cyclic AMP Response Element-Binding Protein/genetics , Gene Expression Regulation, Developmental , Neuromuscular Junction/metabolism , Synapses/physiology , Synaptic Transmission/genetics , Animals , Cyclic AMP Response Element-Binding Protein/metabolism , Drosophila , Excitatory Postsynaptic Potentials/physiology , Homeostasis/genetics , Microscopy, Electron , Motor Neurons/chemistry , Motor Neurons/physiology , Motor Neurons/ultrastructure , Mutation/physiology , Synapses/chemistry , Synapses/ultrastructureABSTRACT
When experimentally challenged with perturbations in synaptic structure and function, neurons have the remarkable ability to regulate their synaptic efficacy back to the normal range. Recent genetic analysis has provided insights into the mechanisms controlling this form of synaptic homeostasis, with implications for our understanding of synaptic development and plasticity.
Subject(s)
Central Nervous System/growth & development , Neuronal Plasticity/genetics , Synapses/physiology , Animals , Central Nervous System/physiology , Homeostasis/physiology , Humans , Neuronal Plasticity/physiologyABSTRACT
Two distinct mechanisms regulate synaptic efficacy at the Drosophila neuromuscular junction (NMJ): a PKA-dependent modulation of quantal size and a retrograde regulation of presynaptic release. Postsynaptic expression of a constitutively active PKA catalytic subunit decreases quantal size, whereas overexpression of a mutant PKA regulatory subunit (inhibiting PKA activity) increases quantal size. Increased PKA activity also decreases the response to direct iontophoresis of glutamate onto postsynaptic receptors. The PKA-dependent modulation of quantal size requires the presence of the muscle-specific glutamate receptor DGluRIIA, since PKA-dependent modulation of quantal size is lost in homozygous viable DGluRIIA- mutants. Furthermore, elevated postsynaptic PKA reduces the quantal amplitude and the time constant of miniature excitatory junctional potential (mEJP) decay to values that are nearly identical to those observed in DGluRIIA mutants. The PKA-dependent reduction in quantal size is accompanied developmentally by an increase in presynaptic quantal content, indicating the presence of a retrograde signal that regulates presynaptic release.
Subject(s)
Cyclic AMP-Dependent Protein Kinases/metabolism , Drosophila/physiology , Presynaptic Terminals/enzymology , Animals , Cyclic AMP-Dependent Protein Kinases/antagonists & inhibitors , Cyclic AMP-Dependent Protein Kinases/genetics , Gene Expression Regulation, Enzymologic , Motor Neurons/enzymology , Mutagenesis , Neuromuscular Junction/enzymology , Neurotransmitter Agents/metabolism , Receptors, Glutamate/genetics , Synaptic Transmission/physiologyABSTRACT
The regulation of synaptic efficacy is essential for the proper functioning of neural circuits. If synaptic gain is set too high or too low, cells are either activated inappropriately or remain silent. There is extra complexity because synapses are not static, but form, retract, expand, strengthen, and weaken throughout life. Homeostatic regulatory mechanisms that control synaptic efficacy presumably exist to ensure that neurons remain functional within a meaningful physiological range. One of the best defined systems for analysis of the mechanisms that regulate synaptic efficacy is the neuromuscular junction. It has been shown, in organisms ranging from insects to humans, that changes in synaptic efficacy are tightly coupled to changes in muscle size during development. It has been proposed that a signal from muscle to motor neuron maintains this coupling. Here we show, by genetically manipulating muscle innervation, that there are two independent mechanisms by which muscle regulates synaptic efficacy at the terminals of single motor neurons. Increased muscle innervation results in a compensatory, target-specific decrease in presynaptic transmitter release, implying a retrograde regulation of presynaptic release. Decreased muscle innervation results in a compensatory increase in quantal size.
Subject(s)
Motor Neurons/physiology , Muscles/physiology , Neuromuscular Junction/physiology , Synapses , Animals , Cell Adhesion Molecules, Neuronal/genetics , Cell Adhesion Molecules, Neuronal/metabolism , Drosophila/embryology , Drosophila/genetics , Drosophila/physiology , Muscles/innervation , MutagenesisABSTRACT
In Drosophila, motoneuron growth cones initially probe many potential muscle targets but later withdraw most of these contacts to form stereotypic synapses with only one or a few muscles. Prior to synapse formation, Fasciclin II (Fas II) is expressed at low levels on muscle. During synapse formation, Fas II concentrates at the synapse and disappears from the rest of the muscle. We previously showed that Fas II is required both pre- and postsynaptically for synaptic stabilization. Here, we show that the differential expression of target-derived Fas II has a profound influence on the patterning of synapse formation. A transient increase in muscle Fas II stabilizes growth cone contacts and leads to novel synapses that are functional and stable; targets that normally receive two inputs can now receive up to six inputs. Changing the relative levels of Fas II on neighboring muscles leads to dramatic shifts in target selection.
Subject(s)
Cell Adhesion Molecules, Neuronal/genetics , Drosophila/genetics , Synapses/chemistry , Synapses/physiology , Animals , Critical Period, Psychological , Drosophila/growth & development , Electrophysiology , Gene Expression Regulation, Developmental/physiology , Genes, Insect/physiology , Larva/genetics , Larva/growth & development , Motor Neurons/chemistry , Motor Neurons/physiology , Motor Neurons/ultrastructure , Muscles/chemistry , Muscles/innervation , Muscles/physiology , Neurites/chemistry , Neurites/physiologyABSTRACT
The glutamatergic neuromuscular synapse in Drosophila forms and differentiates into distinct boutons in the embryo and grows by sprouting new boutons throughout larval life. We demonstrate that two axons form approximately 18 boutons on muscles 7 and 6 by hatching and grow to approximately 180 boutons by third instar. We further show that, after synapse formation, the homophilic cell adhesion molecule Fasciclin II (Fas II) is localized both pre- and postsynaptically where it controls synapse stabilization. In FasII null mutants, synapse formation is normal, but boutons then retract during larval development. Synapse elimination and resulting lethality are rescued by transgenes that drive Fas II expression both pre- and postsynaptically; driving Fas II expression on either side alone is insufficient. Fas II can also control synaptic growth; various FasII alleles lead to either an increase or decrease in sprouting, depending upon the level of Fas II.
Subject(s)
Cell Adhesion Molecules, Neuronal/physiology , Neuromuscular Junction/physiology , Synapses/physiology , Synapses/ultrastructure , Alleles , Animals , Animals, Genetically Modified , Cell Adhesion Molecules, Neuronal/genetics , Drosophila/embryology , Drosophila/genetics , Drosophila/physiology , Embryo, Nonmammalian , Homozygote , Larva , Muscles/innervation , Mutagenesis, Insertional , Neuromuscular Junction/ultrastructure , Neuronal PlasticityABSTRACT
Increased neuronal activity (eag Shaker mutants) and cAMP concentration (dunce mutants) lead to increased synaptic structure and function at the Drosophila neuromuscular junction. Here, we show that the increase in synaptic growth is accompanied by an approximately 50% decrease in synaptic levels of the cell adhesion molecule Fasciclin II (Fas II). This decrease in Fas II is both necessary and sufficient for presynaptic sprouting; FasII mutants that decrease Fas II levels by approximately 50% lead to sprouting similar to eag Shaker and dunce, while transgenes that maintain synaptic Fas II levels suppress sprouting in eag Shaker and dunce. However, FasII mutants that cause a 50% increase in bouton number do not alter synaptic strength; rather, evoked release from single boutons has a reduced quantal content, suggesting that the wild-type amount of release machinery is distributed throughout more boutons.
Subject(s)
Cell Adhesion Molecules, Neuronal/physiology , Neuromuscular Junction/physiology , Neurons/physiology , Synapses/physiology , Synapses/ultrastructure , Animals , Animals, Genetically Modified , Cell Adhesion Molecules, Neuronal/biosynthesis , Cyclic AMP/metabolism , Drosophila , Gene Expression Regulation , Larva , Muscles/innervation , Mutagenesis , Neuromuscular Junction/ultrastructure , Neuronal Plasticity , Neurons/ultrastructureABSTRACT
Increased cAMP (in dunce mutants) leads to an increase in the structure and function of the Drosophila neuromuscular junction. Synaptic Fasciclin II (Fas II) controls this structural plasticity, but does not alter synaptic function. Here, we show that CREB, the cAMP response element-binding protein, acts in parallel with Fas II to cause an increase in synaptic strength. Expression of the CREB repressor (dCREB2-b) in the dunce mutant blocks functional but not structural plasticity. Expression of the CREB activator (dCREB2-a) increases synaptic strength only in FasII mutants that increase bouton number. This CREB-mediated increase in synaptic strength is due to increased presynaptic transmitter release. Expression of dCREB2-a in a FasII mutant background genetically reconstitutes this cAMP-dependent plasticity. Thus, cAMP initiates parallel changes in CREB and Fas II to achieve long-term synaptic enhancement.
Subject(s)
Cyclic AMP Response Element-Binding Protein/metabolism , Drosophila Proteins , Neuromuscular Junction/physiology , Neurons/physiology , Synapses/physiology , Animals , Animals, Genetically Modified , Cell Adhesion Molecules, Neuronal/biosynthesis , Cell Adhesion Molecules, Neuronal/physiology , Cyclic AMP/metabolism , Cyclic AMP Response Element-Binding Protein/biosynthesis , Drosophila , Evoked Potentials , Gene Expression , Larva , Membrane Potentials , Muscle Fibers, Skeletal/physiology , Neuromuscular Junction/ultrastructure , Neuronal Plasticity , Neurons/ultrastructure , Repressor Proteins/biosynthesis , Synapses/ultrastructure , Trans-Activators/biosynthesisABSTRACT
The antimicrobial compound U-82,127 (Pharmacia & Upjohn, Kalamazoo, MI) is a thiopeptide that belongs to a series of cyclic peptide antibiotics produced by Streptomyces arginensis. It is active mainly against Gram-positive organisms. A study involving 576 growing-finishing pigs was conducted at six locations to assess the efficacy of the growth-promoting compound from approximately 19 to 89 kg BW. The basal diet was an unmedicated corn-soybean meal diet fortified with vitamins and trace minerals and containing 16% CP (.80% lysine) during the growing stage (to 54 kg) followed by 13% CP (.60% lysine) during the finishing stage. Dietary dose concentrations of the antimicrobial compound were 0, 3.3, 6.6, and 9.9 mg/kg. At each location, there were six replications of four pigs (two barrows and two gilts) per pen. Diets and water were available for ad libitum consumption. The antimicrobial was provided in coded bags, and investigators were blind to the treatments. The ADG during the growing stage was improved by all levels of the antimicrobial (P < .04), but only the 6.6 mg/kg level improved ADG during the finishing stage (P < .03). Feed:gain was improved by all concentrations of the antibiotic (P < .01) during the growing stage and by the two lower levels of the drug (P < .06) during the finishing stage. Over the entire study, the antimicrobial compound improved ADG (linear, P < .06) and feed:gain (quadratic, P < .01; minimum feed:gain was at 6.2 mg/kg). The lowest dose with a 90% confidence interval of its predicted value not overlapping with the predicted value of the control was 2.3 mg/kg; thus, the efficacious dose range for improving feed/gain was between 2.3 and 6.2 mg/ kg. Neither death loss nor pig removal from the experiment was affected by treatment. The results indicate that the antimicrobial compound U-82,127 is an effective growth-promoting agent for growing-finishing pigs.
Subject(s)
Anti-Bacterial Agents/pharmacology , Growth Substances/pharmacology , Peptides, Cyclic/pharmacology , Swine/growth & development , Animals , Dose-Response Relationship, Drug , Female , Male , Random Allocation , Weight Gain/drug effectsABSTRACT
Since protein kinase C (PKC) has been proven to be a mediator of neutrophil activation and of intracellular calcium homeostasis, its inhibition could protect the myocardium from the deleterious effects of ischemic/reperfusion inury (IRI). The principal objective of this study was to evaluate the efficacy of the PK inhibitor SPC-100270 (2S,3S)-2-amino, 3-octadecanediol in a canine model of IRI. A double-blind study was conducted in which 19 coonhound dogs received either SPC-100270 or a vehicle before going on cardiopulmonary bypass (CPB). After 60 minutes of global normothermic (37 degree C) cardiac arrest (cross-clamp time 65-81 minutes for SPC-100270 and 65-72 minutes for control) and discontinuation of CBP, an epicardial short axis view echocardiogram was performed and reviewed by a double-blinded observer to determine the ejection fraction (EF). EF value exceeded 20% in 5 out of 9 SPC-100270 animals (27%-44%) and in 0 of 10 controls (0%-16%). These data show that SPC-10027 significantly (p=0.01 by Fisher's Exact Test) increased the probability that the animals would exhibit an EF greater than 20%.
Subject(s)
Enzyme Inhibitors/therapeutic use , Myocardial Reperfusion Injury/prevention & control , Protein Kinase C/antagonists & inhibitors , Protein Kinase C/physiology , Sphingosine/analogs & derivatives , Ventricular Function, Left , Animals , Cardiopulmonary Bypass , Dogs , Double-Blind Method , Echocardiography , Electrocardiography , Heart Arrest, Induced , Male , Myocardial Reperfusion Injury/diagnosis , Myocardial Reperfusion Injury/diagnostic imaging , Sphingosine/therapeutic use , Stroke Volume , Time FactorsABSTRACT
Upon contacting its postsynaptic target, a neuronal growth cone transforms into a presynaptic terminal. A membrane component on the growth cone that facilitates synapse formation was identified by means of a complementary DNA-based screen followed by genetic analysis. The late bloomer (lbl) gene in Drosophila encodes a member of the tetraspanin family of cell surface proteins. LBL protein is transiently expressed on motor axons, growth cones, and terminal arbors. In lbl mutant embryos, the growth cone of the RP3 motoneuron contacts its target muscles, but synapse formation is delayed and neighboring motoneurons display an increase in ectopic sprouting.
Subject(s)
Drosophila Proteins , Genes, Insect , Membrane Proteins/physiology , Motor Neurons/physiology , Nerve Tissue Proteins/physiology , Neuromuscular Junction/physiology , Presynaptic Terminals/physiology , Amino Acid Sequence , Animals , Axons/metabolism , Axons/ultrastructure , Cloning, Molecular , Drosophila/embryology , Drosophila/genetics , Drosophila/physiology , Membrane Proteins/chemistry , Membrane Proteins/genetics , Molecular Sequence Data , Motor Neurons/cytology , Motor Neurons/metabolism , Muscles/innervation , Mutation , Nerve Tissue Proteins/chemistry , Nerve Tissue Proteins/genetics , Presynaptic Terminals/ultrastructure , Signal TransductionABSTRACT
Deterioration of tuberculous tissue specimens from badgers during transit in the post was simulated in the laboratory, and it was found that unpreserved tissues showing miliary lesions and initially producing a confluent growth of Mycobacterium bovis became culture-negative over a period of 4 days at ambient summer temperatures (>20 degrees C). These findings implied that false negative results may be obtained from specimens containing only small numbers of bacilli following delays of 2 days between collection and culture. A bench trial of cetyl pyridinium chloride and bromide as preservatives showed viable bacillary numbers were maintained well. A field trial of a 1% aqueous solution of the chloride salt as a transport medium was carried out on 666 split tissue submissions and a significantly improved isolation rate was found from the preserved samples, with 40 (93%) of the total of 43 isolates of M. bovis compared with 27 isolates (63%) when no preservative was used. Isolation using the guinea pig test was attempted on 354 of these samples and yielded 38 (93%) of the 41 isolates, compared with culture of preserved samples which yielded 39 isolates (95%), an insignificant difference. The use of this transport medium maximised cultural isolations, such that no significant gain was achieved by the continued use of the animal test.
ABSTRACT
The dynamics of presynaptic transmitter release are often matched to the physiological properties and function of the postsynaptic cell. Evidence in organisms as diverse as the cricket central nervous system and the cat spinal cord suggests that retrograde signaling is essential for matching presynaptic release properties to the postsynaptic cell. The cricket central nervous system is favorably organized for analysis of synaptic function in the central nervous system. Several lines of independent evidence suggest that it is possible to reliably estimate the size of single quantal release events at the sensory to interneuron synapses of the cricket. A quantal analysis suggests that a retrograde influence on the probability of presynaptic release is responsible for matching presynaptic dynamic properties to postsynaptic targets. This retrograde interaction is hypothesized to be a long-term modification on the basal probability of presynaptic release.
Subject(s)
Neuronal Plasticity/physiology , Neurotransmitter Agents/physiology , Receptors, Presynaptic/physiology , Synapses/physiology , Animals , Cats , Gryllidae , Humans , Neurotransmitter Agents/metabolismABSTRACT
The dynamics of presynaptic transmitter release are often matched to the functional properties of the postsynaptic cell. In organisms ranging from cats to crickets, evidence suggests that retrograde signaling is essential for matching these presynaptic release properties to individual postsynaptic partners. Retrograde interactions appear to control the development of presynaptic, short-term facilitation and depression.