Acidic amino acid binding sites in mammalian neuronal membranes: their characteristics and relationship to synaptic receptors

Glutamate’s role as a neurotransmitter at synapses has been known for 40 years, but glutamate has since been shown to regulate neurogenesis, neurite outgrowth, synaptogenesis and neuron survival in the developing and adult mammalian nervous system. Cell surface glutamate receptors are coupled to Ca2+ influx and release from endoplasmic reticulum stores which causes rapid (kinase- and protease-mediated) and delayed (transcription-dependent) responses that change the structure and function of neurons. Neurotrophic factors and glutamate interact to regulate developmental and adult neuroplasticity. For example, glutamate stimulates the production of brain-derived neurotrophic factor (BDNF) which, in turn, modifies neuronal glutamate sensitivity, Ca2+ homeostasis and plasticity. Neurotrophic factors may modify glutamate signalling directly, by changing the expression of glutamate receptor subunits and Ca2+-regulating proteins, and also indirectly by inducing the production of antioxidant enzymes, energy-regulating proteins and anti-apoptotic Bcl2 family members. Excessive activation of glutamate receptors, under conditions of oxidative and metabolic stress, may contribute to neuronal dysfunction and degeneration in diseases ranging from stroke and Alzheimer’s disease to psychiatric disorders. By enhancing neurotrophic factor signalling, environmental factors such as exercise and dietary energy restriction, and chemicals such as antidepressants may optimize glutamatergic signalling and protect against neurological disorders.

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Glutamate and neurotrophic factors in neuronal plasticity and disease.

Transient and enduring morphological correlates of synaptic activity and efficacy change in the rat hippocampal slice.

1. A study was made of functional and structural changes during degeneration of end-plates in the rat diaphragm after phrenic nerve section at two levels.



2. For 8–10 hr after cutting the nerve in the neck, all end-plates retain the ability to transmit impulses. During the following 8–10 hr, an increasing number of end-plates lose this ability so that after a total of about 20 hr, no end-plates can transmit.



3. Transmission failure occurs abruptly at most end-plates. This failure is usually accompanied by cessation of spontaneous miniature end-plate potentials (min.e.p.p.s), though in a few cases min.e.p.p.s persist after junctional transmission has failed. Several degenerating junctions were observed where the frequency of min.e.p.p.s was very low, suggesting an intermediate stage in min.e.p.p. failure.



4. The time of junctional failure depends on the length of the degenerating nerve stump. For each additional centimetre of nerve, failure is delayed about 45 min.



5. Changes in ultrastructure of nerve endings closely parallel those of function. For about 8–12 hr after cutting the nerve, nearly all end-plates appear normal. During the period when transmission is failing, some end-plates are clearly undergoing structural break-down. By the time functional failure is complete, all end-plates appear grossly abnormal.



6. During degeneration, the contents of the axoplasm undergo disruption and the nerve terminal breaks up into small fragments. In contrast, the Schwann cell appears to become very active and its processes extend into the synaptic cleft to surround fragments of the nerve terminal. Ultimately, the Schwann cell completely replaces the axon at the end-plate.



7. Increasing the length of the peripheral nerve stump delays the onset of structural break-down. Disruption of end-plates near the site of nerve entry into the muscle occurs before those farther away.



8. It is suggested that end-plate degeneration is triggered by a signal which passes from the site of injury to the nerve terminal. The duration of the period after transection when end-plates appear to be normal would then reflect the time required for this signal to travel the length of the isolated nerve stump.

https://physoc.onlinelibrary.wiley.com/doi/pdf/10.1113/jphysiol.1970.sp009076

On the degeneration of rat neuromuscular junctions after nerve section

The possibility that L-glutamate is the excitatory transmitter at the Drosophila larval neuromuscular junction and the ionic basis of its action on the muscle membrane are examined. 2. Iontophoretically applied L-glutamate causes muscle depolarization (L-glutamate potential) if and only if the L-glutamate pipette is within a few mum of the nerve ending. D-glutamate, substance P, ACh and GABA are ineffective. 3. Bath-applied L-glutamate produces similar changes in the time course and amplitude of miniature excitatory junctional potential (m.e.j.p.), excitatory junctional potential (e.j.p.) and the L-glutamate potential. 4. Neuromuscular transmission and excitation-contraction coupling are operative in a haemolymph-like solution containing 1 mM L-glutamate. 5. The reversal potentials of the e.j.p. and the L-glutamate potential are identical to each other, changing similarly with changes in the ionic compositions of the external medium (twelve solutions). 6. The ionic dependence of the reversal potentials is predicted from an extended constant-field equation using a ratio of sodium:potassium permeabilities of PNa/PK=1-3, and a ratio of magnesium:potassium permeabilities of PMg/PK=4-7. 7. It is concluded that L-glutamate is, or is an agonist of, the excitatory transmitter at certain Drosophila larval neuromuscular junctions.

L-glutamate as an excitatory transmitter at the Drosophila larval neuromuscular junction.

The presence of extrajunctional acetylcholine (ACh) receptors on vertebrate skeletal muscle fibres is well established1, 2. Fast muscle fibres in the rat possess extrajunctional receptors only in the vicinity of neuromuscular junctions while slow fibres exhibit a measurable ACh sensitivity over the entire non-synaptic membrane with peaks of sensitivity at the nerve-muscle and muscle-tendon junctions3. Sodium and potassium ions are involved in the conductance increase seen during activation of both junctional and extrajunctional ACh receptors on frog muscle fibres, but the relative contribution of potassium is greater in the case of the extrajunctional receptors4.

Two populations of L-glutamate receptors on locust muscle fibres.

GLUTAMIC acid mimics the transmitter at the crustacean excitatory neuromuscular synapse1 and has been tentatively identified with the chemical mediator at the insect peripheral excitatory synapse2,3. Similarities in the pharmacology of insect and crustacean excitatory neuromuscular synapses are, perhaps, to be expected since the pharmacological properties of their peripheral inhibitory synapses are remarkably similar2,4. In an effort to determine the site of action of glutamic acid on the insect neuromuscular system we have examined the effects of this substance on the electrical and mechanical properties of a locust neuromuscular preparation.

Chemical Transmission at the Insect Excitatory Neuromuscular Synapse

THE release of acetylcholine from vertebrate motor nerve terminals has been demonstrated several times since the original observation by Dale and his colleagues in the 1930s, and identification of acetylcholine as the transmitter at vertebrate nerve–muscle synapses now seems well established. At locust and grasshopper muscle synapses, however, where acetylcholine has no pharmacological action1,2, it has been suggested that L-glutamate mediates transmission at excitatory synapses3–5 and γ-aminobutyric acid mediates transmission at inhibitory synapses3,6. This communication reports studies on the release of glutamate from endings of peripheral excitatory axons (motor axons) in the locust, Schistocerca gregaria. The effects of iontophoretic application of L-glutamate to locust muscle fibres have also been studied in an attempt to pinpoint the site or sites of action of this substance.

L-Glutamate at insect excitatory nerve-muscle synapses.

L-GLUTAMATE, rather than acetylcholine, is probably the transmitter at the excitatory synapses found on insect skeletal muscle fibres1–8. By applying L-glutamate iontophoretically from a micropipette to localized areas on the surface of locust muscle fibres the glutamate sensitivity of the muscle membrane is found to decrease sharply with distance from a synaptic site5,7,8. Within 10–35 microns of a synapse the glutamate sensitivity falls either to zero or to a low background level about 10,000 times less than that at the synaptic sites5. Insect skeletal muscle fibres are multiterminally innervated and the synapses are usually on the inner faces of the fibres. The areas of fibre membrane most sensitive to glutamate can therefore be most frequently located by positioning the glutamate electrode in a cleft between two fibres (Fig. 1b), Usually the outer, exposed surfaces of the fibres are devoid of synaptic membrane and are either insensitive to glutamate or sensitive to only very high concentrations of this amino-acid (Fig. 2). Here I report studies of denervated locust muscle fibres, which demonstrate that, following denervation, the glutamate receptors can be found over the entire surface of the muscle fibres.

Glutamate Sensitivity of Denervated Insect Muscle Fibres

THERE is considerable evidence that L-glutamate is the excitatory transmitter both at the insect and crustacean neuromuscular junction1,2, and also in the vertebrate central nervous system3. Here we report the use of the voltage clamp technique to compare the reversal potentials of the conductance increase produced by the excitatory transmitter and the application of L-glutamate at the insect neuromuscular junction. We also investigated the ionic basis of the L-glutamate conductance increase using voltage clamping. These experiments could be carried out accurately because changes in resistance of the electrically excitable membrane caused by changes in membrane potential or by different ionic media do not affect the amplitude of the excitatory junctional currents or glutamate currents.

P. N. R. Usherwood

Papers

Two populations of L-glutamate receptors on locust muscle fibres.

Chemical Transmission at the Insect Excitatory Neuromuscular Synapse

L-Glutamate at insect excitatory nerve-muscle synapses.

Glutamate Sensitivity of Denervated Insect Muscle Fibres

Voltage clamp studies of glutamate synapse