A.J. Anderson

Characterization of a potassium channel toxin from the Caribbean sea anemone Stichodactyla helianthus

A peptide toxin, ShK, that blocks voltage-dependent potassium channels was isolated from the whole body extract of the Caribbean sea anemone Stichodactyla helianthus. It competes with dendrotoxin I and alpha-dendrotoxin for binding to synaptosomal membranes of rat brain, facilities acetylcholine release at an avian neuromuscular junction and suppresses K+ currents in rat dorsal root ganglion neurones in culture. Its amino acid sequence is R1SCIDTIPKS10RCTAFQCKHS20MKYRLSFCRK30TCGTC35. There is no homology with other K+ channel-blocking peptides, except for BgK from the sea anemone Bunodosoma granulifera. ShK and BgK appear to be in a different structural class from other toxins affecting K+ channels.

Dendrotoxins: Snake toxins that block potassium channels and facilitate neurotransmitter release

This scientific review looks at dendrotoxins: snake toxins that block potassium channels and facilitate neurotransmitter release

A potassium channel toxin from the secretion of the sea anemone Bunodosoma granulifera. Isolation, amino acid sequence and biological activity.

A peptide toxin affecting potassium channels was isolated from the sea anemone Bunodosoma granulifera. It facilitates acetylcholine release at avian neuromuscular junctions, competes with dendrotoxin I, a probe for voltage-dependent potassium channels, for binding to synaptosomal membranes of rat brain with a Ki of 0.7 nM and suppresses K+ currents in rat dorsal root ganglion neurones in culture. It represents a new structural type of potassium channel toxin with the sequence V1RCDWFKETA10CRHAKSLGNC20RTSQKYRANC30AKTLQCC37 (M(r) 4275, three disulfides).

Effects of the potassium channel blocking dendrotoxins on acetylcholine release and motor nerve terminal activity

1 The effects of the K+ channel blocking toxins, the dendrotoxins, on neuromuscular transmission and motor nerve terminal activity were assessed on frog cutaneous pectoris, mouse diaphragm and mouse triangularis sterni nerve‐muscle preparations. Endplate potentials (e.p.ps) and miniature e.p.ps were recorded with intracellular microelectrodes, and nerve terminal spikes were recorded with extracellular electrodes placed in the perineural sheaths of motor nerves. 2 Dendrotoxin from green mamba (Dendroaspis angusticeps) venom and toxin I from black mamba (D. polylepis) venom increased the amplitude of e.p.ps by increasing quantal content, and also induced repetitive e.p.ps. 3 Perineural recordings revealed that dendrotoxins could decrease the component of the waveform associated with K+ currents at the nerve terminals, and induce repetitive activation of nerve terminals. 4 In frog motor nerves, dendrotoxins are known to block the fast f1 component of the K+ current at nodes of Ranvier. Blockade of a similar component of the K+ current at motor nerve terminals may be responsible for the effects of these toxins on neuromuscular transmission. 5 Similar conclusions can be drawn from the results obtained from mouse neuromuscular junctions.

Omega-conotoxin does not block the verapamil-sensitive calcium channels at mouse motor nerve terminals.

Release of acetylcholine at the neuromuscular junctions of skeletal muscle is not sensitive to organic Ca2+ channel blockers. However, in mouse motor nerve endings, extracellular recording reveals that a verapamil-sensitive Ca2+ current can be induced after block of K+ channels. Recordings of extracellular action potentials from inside the perineural sheaths of nerves innervating mouse triangularis sterni muscles reveal that this verapamil-sensitive current is not blocked by omega-conotoxin, and hence, it does not involve channels similar to the L-channels of neuronal cell bodies.

Effects of charybdotoxin, a blocker of Ca2+ ‐activated K + channels, on motor nerve terminals

1 The contribution of Ca2+‐activated K+ currents (IK,Ca)to the control of electrical excitability of motor nerve terminals and the control of acetylcholine release was assessed by studying the effects of the specific K(Ca) channel blocking toxins charybdotoxin and apamin. Electrical activity of the terminal regions of motor nerves was assessed by extracellular recording from an electrode placed in the perineural sheaths of nerves in the mouse triangularis sterni and frog cutaneous pectoris preparations. Acetylcholine release was monitored by intracellular recording of endplate potentials (e.p.ps). 2 Charybdotoxin (20–300 nm), but not apamin (10nM‐2.5 μm), selectively reduced the amplitude of an IK,Ca unmasked by prior blockade of the delayed rectifier K+ current with 3,4‐diaminopyridine (3,4‐DAP). 3 In the combined presence of 3,4‐DAP and charybdotoxin, large Ca2+‐dependent plateau responses developed, but only moderate and transient increases in acetylcholine release occurred. 4 In the absence of 3,4‐DAP, charybdotoxin did not alter the electrical activity of, or the transmitter release from motor nerve terminals. 5 A possible role of the charybdotoxin‐sensitive IK,Ca in the control of transmitter release is discussed.

Toxins from Mamba Venoms that Facilitate Neuroiluscular Transmission

AbstractVenoms from the four species of African mambas have many neurotoxins that are different from other snake neurotoxins. Postjunctional α-neurotoxins, which bind to nicotinic cholinoceptors, appear to be the only type of toxin that mambas have in common with other snakes with neurotoxic venoms. Typical mamba neurotoxins are prejunctional facilitatory toxins and anticholinesterase toxins or fasciculins.The prejunctional facilitatory toxins enhance the amount of transmitter that is released in response to nerve stimulation. This activity was first demonstrated with dendrotoxin (from Dendroaspis anqusticeps) at the neuromuscular junction, but it can also be observed in both the sympathetic and parasympathetic branches of the autonomic nervous system and in the central nervous system. These facilitatory neurotoxins have 57-60 amino acids in a single polypeptide chain cross-linked by three disulphide bonds. They are structurally homologous to the Kunitz type protease inhibitors such as bovine pancreatic t...

Apparent block of K+ currents in mouse motor nerve terminals by tetrodotoxin, μ-conotoxin and reduced external sodium

1 In mouse triangularis sterni nerve‐muscle preparations, reduced extracellular Na+ concentrations and low concentrations of the Na+ channel blocking toxins tetrodotoxin (TTX, 18–36 nm) and μ‐conotoxin GIIIB (0.4–2.0 μm) selectively decreased the amplitude of the component of perineural waveforms associated with nerve terminal K+ currents, without affecting the main Na+ spike. 2 Intracellular recording of endplate potentials (e.p.ps) and miniature endplate potentials (m.e.p.ps) from triangularis sterni preparations revealed that TTX and μ‐conotoxin GIIIB depressed the evoked quantal release of acetylcholine without significant effects on m.e.p.p. amplitude, frequency or time constant of decay. 3 The apparent block of K+ current by low concentrations of TTX and μ‐conotoxin is probably not a direct effect on K+ channels but results from a decrease in the passive depolarization of nerve terminals following blockade of a small proportion of axonal Na+ channels.

Effects of the facilitatory compounds catechol, guanidine, noradrenaline and phencyclidine on presynaptic currents of mouse motor nerve terminals

SummaryCatechol, guanidine, noradrenaline, and phencyclidine can increase acetylcholine release at neuromuscular junctions. To determine if they act by affecting nerve terminal action potentials, the electrical activity of the terminal regions of motor nerves was recorded with an extracellular electrode inserted in the perineural sheaths of nerves in the mouse triangularis sterni preparation. Catechol (from 10 μM) and guanidine (from 1 mM) produced a selective reduction in the component of the perineural waveform associated with voltage-dependent K+ currents, without significant effects on Na+, Ca+, or Ca2+-activated K+ currents. A selective block of K+ channels in nerve terminals would cause a prolonged depolarization and hence a large influx of Ca2+ to trigger acetylcholine release; this could explain the facilitatory effects of guanidine and catechol. Noradrenaline produced a slight increase in the amplitude of the. perineural waveform. This is consistent with hyperpolarization of the resting membrane potential of the nerve, which could lead to facilitation of acetylcholine release. Phencyclidine blocked Na+- and K+-related portions of the signal.

Effects of fasciculin 2, an anticholinesterase polypeptide from green mamba venom, on neuromuscular transmission in mouse diaphgragm preparations

Fasciculin 2, a polypeptide from green mamba (Dendroaspis angusticeps) venom, causes an increase in the twitch response of mouse phrenic nerve-hemidiaphragm preparations to indirect stimulation. Intracellular recording reveals that fasciculin 2 augments neuromuscular transmission by increasing the amplitude and duration of endplate potentials. Its action is not reversed by washing. Interactions with neostigmine confirm that fasciculin 2 acts as an anticholinesterase. It has no presynaptic actions on transmitter release or postsynaptic receptor blocking actions. On chicken muscle preparations, fasciculin 2 has no anticholinesterase actions. Because of this selectivity and its apparent irreversibility, fasciculin 2 should be useful in characterizing different forms of acetylcholinesterase.