Block of an insect central nervous system GABA receptor by cyclodiene and cyclohexane insecticides

The effects of a cyclodiene (endrin) and a cyclohexane (lindane) insecticide have been tested on gamma-aminobutyric acid (GABA) receptors in the central nervous system of the cockroach (Periplaneta americana), by using electrophysiological methods and an in vitro functional receptor assay. In electrophysiological experiments on an identified motor neuron (Df), endrin blocked the GABA response with a 50% inhibition concentration of 5.0 x 10(-7) M in a non-competitive manner. The actions of endrin were irreversible under the experimental conditions adopted. Increasing the intracellular chloride concentration reduced the effectiveness of endrin, whereas a change in the potassium concentration failed to influence the block by endrin of GABA responses. Lindane exhibited similar actions to endrin on insect GABA receptors, but was approximately an order of magnitude less effective. In a microsac preparation from cockroach nerve cords, endrin, at a concentration of 1.0 x 10(-5) M, completely blocked GABA-stimulated 36Cl- uptake, whereas the same concentration of lindane was less potent, only blocking about 40% of uptake under similar conditions. Neither insecticide had any effect on L-glutamate-activated chloride channels. The results demonstrate that endrin and lindane block functional insect neuronal GABA receptors.     

Glial repair in the cultured central nervous system of an insect

Summary Insect glial cells are capable of division and repair in organ culture after selective damage with the toxin ethidium bromide. The repair is slower and less organised than seen in vivo after similar treatment and is still incomplete after one month. Granule-containing cells, which play an important role in the early stages of repair in vivo, are never seen in cultured connectives. This observation adds further support to the hypothesis that these cells are derived from haemocytes and that their presence is necessary for rapid and orderly repair. The uptake of ³H-thymidine into perineurial glial cells in vitro, both in control and ethidiumtreated connectives, shows that there is a considerable proliferation of cells in this region. Some uptake of thymidine is also seen in subperineurial glia but division alone cannot account for the large increase in the number of glial nuclei found at the early stages of repair in this region. Further, glial cells with diverse morphologies suggest that subpopulations are present. We conclude that cell migration from undamaged areas, as well as cell proliferation, is necessary for CNS repair in vitro.     

Modulatory actions of norepinephrine in the central nervous system.

Early studies have shown that norepinephrine (NE) released synaptically or iontophoretically onto neurons in the central nervous system acts to depress firing by a mechanism associated with a hyperpolarization but no change or an increase in membrane resistance. This in contrast to classical transmitters, which cause hyperpolarization by a conductance increase. Recent studies designed to clarify the functional implications of these biophysical actions have revealed new phenomenons in which the major overall effect of NE on cerebellar Purkinje cells is to enhance conventional synaptic input and induce an increase in signal-to-noise ratio of evoked versus spontaneous activity. NE released iontophoretically or via stimulation of the locus coeruleus also has been found to enhance the inhibitory effects of gamma-aminobutyric acid, an endogenous cerebellar transmitter. The effects appear even at low doses of NE having no direct depressant action on spontaneous activity. Specificity tests have shown no enhancement of glycine-induced inhibition by NE and an inability of dopamine to mimic NE. The hypothesis is presented that a significant action of NE in the central nervous system is to induce a bias that alters postsynaptic responsiveness to conventional transmitter systems, which themselves may be more directly concerned with detailed information transfer.     

Taurine in the insect central nervous system

1. Taurine is one of the most abundant free amino acids found in the tissues of insect nervous systems. A brief survey of its immunocytochemical distribution is provided for the brain of worker honeybees.2. The protocerebral mushroom bodies are prominent neuropiles of the insect brain. Immunoreactivity for taurine was compared in the mushroom body intrinsic Kenyon cells of Apis, Drosophila, and Locusta.3. In all three species Kenyon cells expressed immunoreactivity.4. The intensity of the immunoreactivity was, however, graded, depending on the species.5. Recent technical advances in the primary culture of the Kenyon cells of honeybees in a defined taurine-free medium provide the opportunity to investigate the action of taurine in a controlled environment.6. Taurine-like immunoreactivity has been described in the photoreceptor cells of insect and mammalian visual systems. Physiological evidence for similar functions of taurine in mammalian and insect nervous systems is reviewed.     

Proctolin potentiates synaptic transmission in the central nervous system of an insect

1. Bursts of spike activity in the ventral nerve cord of the cockroach were elicited by mechanically stimulating the cercal organs. 2. In the presence of micromolar proctolin, the peak frequency and the duration of a burst were slowly but significantly increased. 3. In contrast, carbachol produced an immediate enhancement of spontaneous activity, but a potentiation of bursts was not seen. 4. It is proposed that proctolin functions as a neuromodulator in the terminal abdominal ganglion of the cockroach.     

Enzyme effects on the connective tissues of an insect central nervous system

The effect of various enzymes on the two connective tissue matrices of the cockroach central nervous system were investigated. Removal of the neural lamella, using collagenase, allows some of the cells which form the perineurium to pull out of this cell layer but the perineurial bracelet cells maintain an intact blood-brain barrier. Incubation of the nerve cord with hyaluronidase has little or no effect on the neural lamella and allows the selective removal of the matrix from the glial lacunar system. Partial removal of this matrix appears to have little effect on the ability of the axons to conduct action potentials at high frequencies. In addition to this difference in susceptibility of the neural lamella and lacunar matrices to different enzymes, there appears to be a difference between the lacunar matrix of the connectives and of the ganglia, the latter being more resistant to enzyme attack. There is no such difference in the neural lamella covering the ganglia and connectives.     

Degeneration and regeneration in the insect central nervous system. I

Summary After cutting a neck connective of Schistocerca gregaria, only 2% of the axons on each side of the lesion degenerate. The remainder show reactive changes, which last for approximately one week at 28° C. There is no morphological change in either of the pro/mesothoracic connectives after injury to the neck connective. Phagocytes invade the stumps, but attack only degenerating cells, and are absent by Day 7.Regeneration from the connective stumps begins a week after injury; a functional link may be formed by Day 10, but by Day 23 the new connective cannot function adequately for the locust's survival, if the undamaged connective is then cut.The chief morphological changes in the reactive axoplasm are increases in the number of mitochondria, neurotubules, vesicles and vacuoles. These changes appear to be a local response, and not to be influenced by the neuron cell bodies. Some glial cytoplasm (presumably enucleated), degenerates rapidly after injury, and replacement begins by Day 5. Tracheoles, never seen in normal connectives appear in the reactive connective from Days 3–8, this is interpreted as a migration from the ganglion in response to oxygen deficiency in the connective.The results are discussed in relationship to previous work.This work was supported by a Study and Serve grant from the British Government, and a grant from the Worshipful Company of Goldsmiths.I wish to acknowledge the help and advice given to me by Dr. C. H. F. Rowell.     

c-Fos-related antigens in the central nervous system of an insect, Acheta domesticus

Fos-related antigens (Fra) were detected in the nuclei of neurones in young adult Acheta domesticus female crickets by immunohistochemical analysis, using an antibody that recognizes the amino-acid sequence 127-152 of c-Fos protein. Specificity of Fra immunoreactivity was confirmed by Western blot analysis of nuclear extracts from neural tissues. A major immunoreactive doublet with an apparent molecular mass of 52,000/54,000 Da was detected in nuclear extracts. Immunostaining of the 52,000/54,000 Da doublet showed variations in intensity during the first 5 days following the imaginal molt. Staining was more intense between day 2 and day 4 when ecdysteroid titers were high. Expression of Fra was low in allatectomized (i.e., deprived of juvenile hormone and ecdysteroids) and ovariectomized (i.e., deprived of ecdysteroids) females as compared to control females. These results show the involvement of hormone-regulated process in expression of Fra. The effect of nociceptive stimulation on Fra expression was tested. Twenty minutes after removal of the ovipositor, a supplementary band with an apparent molecular mass of 70,000 Da appeared in the nuclear extracts, then decreased and disappeared totally after 45 min. Several other Fos-related antigens with different temporal patterns of expression were also detected.     

Receptors for L-glutamate and GABA in the nervous system of an insect (Periplaneta americana)

The nervous system of the cockroach Periplaneta americana is well suited to studies of invertebrate amino acid receptors. Using a combination of radioligand binding and electrophysiological techniques, several distinct receptors have now been identified. These include an l-glutamate-gated chloride channel which has no known counterpart in the vertebrate nervous system, and a putative kainate/quisqualate receptor with pharmacological properties different from those of the existing categories of vertebrate excitatory amino acid receptors. GABA receptors have also been characterized in the cockroach nervous system. Bicuculline, benzodiazepines and steroids have revealed important differences between certain insect GABA-gated chloride channels and vertebrate GABA receptors. Identifiable neurones may facilitate the allocation of specific functions to amino acid receptor subtypes. In view of the existence of subtypes of amino acid receptors in insects, it is of interest to examine how this is reflected at the molecular level in terms of receptor subunit composition and amino acid sequence. Preliminary molecular cloning studies on insect GABA receptors are described.     

The electrophysiological actions of neurotensin in the central nervous system

The endogenous neuropeptide, neurotensin (NT) alters the firing frequencies of certain neurons in the central nervous system (CNS). This is one of the findings that support the hypothesis that NT is a neurotransmitter substance. The direct application of NT on CNS neurons causes predominantly excitatory effects. These effects occur in a dose-related fashion via a calcium-dependent postsynaptic mechanism. The C-terminal hexapeptide fragment, NT 8-13 exerts similar electrophysiological effects to NT, while the N-terminal octapeptide fragment, NT 1-8 is devoid of such activity. NT produces a significant increase in the firing rates of individual neurons in the substantia nigra (SN), ventral tegmental area (VTA), medial prefrontal cortex (MPF), hypothalamus, and periaqueductal grey (PAG). This excitation occurs with a rapid onset and is readily reversible after cessation of NT application. In contrast, NT has no effect or weak inhibitory effects on the firing rates of neurons in the locus coeruleus (LC) and cerebellum. These electrophysiological actions of NT appear to be unique and not shared by other neurotransmitter and neuropeptide receptor antagonists and agonists that have been studied via direct co-application. NT attenuates dopamine (DA)-induced inhibition associated with direct application onto neurons in the SN and VTA both in vivo and in vitro. Intracellular recordings suggest that direct application of higher concentrations of NT appears to produce 'depolarization block' on individual neurons in the SN, VTA, MPF, and hypothalamus. The electrophysiological consequences of NT application not only show similarities to clinically efficacious antipsychotic medications, but also demonstrate the ability of NT to modulate the activity of dopamine (DA) neurons at the cellular level via specific NT binding sites. These findings further underscore the possibility that NT may play a pre-eminent role in the pathogenesis of, and psychopharmacological management of neurological and psychiatric disorders purportedly related to perturbation of CNS DA systems including schizophrenia.     

Characterization of the synaptic actions of an interneuron in the central nervous system of Tritonia

The motor program that drives the swimming behavior of the marine mollusk Tritonia diomedea is generated by three interneuronal populations in the cerebral ganglia. One of these populations, the pair of C2 neurons, is shown to also exert powerful synaptic actions upon most cells in the contralateral pedal ganglion. Intracellular staining with Co²⁺ showed that the C2 neurons projected to the contralateral pedal ganglion as a single unbranched axon, and nearly all contralateral pedal neurons received monosynaptic input from C2. Orthodromic stimulation of most peripheral nerves caused monosynaptic excitation of C2 by afferent sensory cells and, in some cases, monosynaptic inhibition from an unidentified source. C2 neurons produced four types of postsynaptic potential (PSP) on pedal neurons: (1) a fast, Cl^?-mediated inhibition (FIPSP); (2) a fast, Na⁺-mediated excitation (FEPSP); (3) a slow, K⁺-mediated inhibition (SIPSP); and (4) a slow, conductance-decrease excitation (SEPSP). All four could be recorded simultaneously in some pedal neurons. The C2 neurons appear to be high-order, multiaction neurons involved in both the generation of a complex motor program and the coordination of ancillary neuronal activity.     

Immunocytochemical localization of sodium channels in an insect central nervous system using a site-directed antibody

Antibodies to channel proteins and specific peptide sequences have been previously used to localize voltage-activated sodium channels in the rat brain. Here we describe the first localization of sodium channels in an insect nervous system using a site-directed antibody. The mesothoracic ganglion of the cockroach was stained with an antibody to the highly conserved SP19 sequence. Antibody labelling was visualized by light microscopy using the avidin/biotin method on was sections, and transmission electron microscopy of immunogold-labelled thin sections. Central ganglia of insects contain clearly separated regions of cell bodies, synaptic neuropil, axon tracts, and nerves. Antibody staining by light microscopy was limited to neurons, and was intense in axons throughout the ganglion and nerves. Staining was also strong in the cytoplasm, but not the nuclei, of many neuronal cell bodies. Neuropil regions were relatively lightly labelled. These findings can be correlated with the known electrophysiology of the ganglion. Electron microscopy detected sodium channels in areas surrounding axons, probably including axon membranes and enveloping glial cell membranes. Axonal mitochondria were also heavily labelled, suggesting a sodium channel transport function for these organelles. © 1993 John Wiley & Sons, Inc.     

Serotonin immunoreactive neurons in the central nervous system of an insect (Periplaneta americana)

Serotonin-like immunoreactivity was mapped in the central nervous system (CNS) of the cockroach, Periplaneta americana. Immunoreactive staining occurred in every ganglion of the CNS. The largest numbers of immunoreactive somata were detected in the optic lobes and the brain, and lowest numbers in the first and second thoracic ganglia. Dense stained fibers, ramifications, and varicosities were found in all ganglia, and numerous axon like processes occurred in all interganglionic connectives. Immunoreactive processes were not, however, detected in most of the peripherally projecting nerve roots. Processes were found only in roots of the suboesophageal ganglion and the tritocerebral lobes of the brain. A comparison of the map for serotonin immunoreactivity with one generated for the pentapeptide transmitter proctolin suggests that the two systems overlap only in the suboesophageal ganglion and the tritocerebrum. The amine and peptide may co-occur in neurons in these regions. The serotonin immunoreactive system appeared significantly different from the octopaminergic system of the ventral nerve cord. Seventy-two potentially identifiable immunoreactive cells were located in the cockroach CNS. Some of these may be suitable for physiological study of the functional role of serotonin.     

The origins of cell diversity in the insect central nervous system

There are thousands of unique neurons and many types of glia in the insect central nervous system. How is this cell diversity generated? Neurogenesis begins with the delamination and enlargement of individual cells of the ventral ectoderm to form a stereotyped array of neuroblasts. Every neuroblast divides asymmetrically to generate a chain of approximately 10 smaller progeny, each of which produces a pair of neurons. Ablation, transplantation and in vitro culture experiments illuminate the role of cell interactions and cell lineage during neurogenesis, and genetic approaches in Drosophila are beginning to provide insight into the molecular mechanisms controlling these events.     

The neurotoxic actions of quinolinic acid in the central nervous system

Excitotoxins such as kainic acid, ibotenic acid, and quinolinic acid are a group of molecules structurally related to glutamate or aspartate. They are capable of exciting neurons and producing axon sparing neuronal degeneration. Quinolinic acid (QUIN), an endogenous metabolite of the amino acid, tryptophan, has been detected in brain and its concentration increases with age. The content of QUIN in the brain and the activity of the enzymes involved in its synthesis and metabolism show a regional distribution. The neuroexcitatory action of QUIN is antagonized by magnesium (Mg2+) and the aminophosphonates, proposed N-methyl-D-aspartate (NMDA) receptor antagonists, suggesting that QUIN acts at the Mg2+ -sensitive NMDA receptor. Like its excitatory effects, QUIN's neurotoxic actions in the striatum are antagonized by the aminophosphonates. This suggests that QUIN neurotoxicity involves the NMDA receptor and (or) another receptor sensitive to the aminophosphonates. The neuroexcitatory and neurotoxic effects of QUIN are antagonized by kynurenic acid (KYN), another metabolite of tryptophan. QUIN toxicity is dependent on excitatory amino acid afferents and shows a regional variation in the brain. Local injection of QUIN into the nucleus basalis magnocellularis (NBM) results in a dose-dependent reduction in cortical cholinergic markers including the evoked release of acetylcholine. A significant reduction in cortical cholinergic function is maintained over a 3-month period. Coinjection of an equimolar ratio of QUIN and KYN into the NBM results in complete protection against QUIN-induced neurodegeneration and decreases in cortical cholinergic markers. In contrast, focal injections of QUIN into the frontoparietal cortex do not alter cortical cholinergic function.(ABSTRACT TRUNCATED AT 250 WORDS)     

Possible involvement of GABA in the central actions of benzodiazepines.

The effects of several benzodiazepines on a variety of nervous activities known or presumed to depend on GABA are presented and compared with those of agents that deplete or increase the level of endogenous GABA: antagonism of various convulsant agents in mice, enhancement of presynaptic inhibition in the spinal cord and the cuneate nucleus of cats, decrease of the spontaneous firing rate of cerebellar Purkinje cells in cats and rats, antagonism of bicuculine-induced depression of the strio-nigral-evoked potential in the cat, potentiation of haloperidol-induced catalepsy in rats, GABA-mimetic actions on drug-induced PGO-waves in cats and on eserine-induced circling in guinea pigs. Diazepam slightly increased the GABA level in the cat spinal cord and in the total brain of mice and rats; this increase does not seem to be due to an increase of GABA synthesis. It is concluded that benzodiazepines probably enhance presynaptic inhibition at all levels of the neuraxis and that this effect requires not only the presence of GABA but is also dependent on an activity of GABA-ergic neurons. Benzodiazepines also appear to enhance postsynaptic inhibition where this is mediated by GABA. Many actions of benzodiazepines can be tentatively explained by a stimulus-bound enhancement of GABA effects.