Pyramidal neurons receive tens of thousands of synaptic inputs on their dendrites. The dendrites dynamically alter the strengths of these synapses and coordinate them to produce an output in ways that are not well understood. Surprisingly, there turns out to be a very high density of transient A-type potassium ion channels in dendrites of hippocampal CA1 pyramidal neurons. These channels prevent initiation of an action potential in the dendrites, limit the back-propagation of action potentials into the dendrites, and reduce excitatory synaptic events. The channels act to prevent large, rapid dendritic depolarizations, thereby regulating orthograde and retrograde propagation of dendritic potentials.

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K + channel regulation of signal propagation in dendrites of hippocampal pyramidal neurons

Diversity and ubiquity of K channels.

On the basis of electrophysiological analysis of Shaker mutants, the Shaker locus of Drosophila melanogaster has been proposed to encode a structural component of a voltage-dependent potassium channel, the A channel. Unlike sodium channels, acetylcholine receptors, and calcium channels, K+ channels have not been purified biochemically. To facilitate biochemical studies of a K+ channel, genomic DNA from the Shaker locus has been cloned. Rearrangements in five Shaker mutants have been mapped to a 60-kilobase segment of the genome. Four complementary DNA clones have been analyzed. These clones indicate that the Shaker gene contains multiple exons distributed over at least 65 kilobases of genomic DNA in the region where the mutations mapped. Furthermore, the gene may produce several classes of alternatively spliced transcripts. Two of the complementary DNA clones have been sequenced and their sequences support the hypothesis that Shaker encodes a component of a K+ channel.

https://science.sciencemag.org/content/sci/237/4816/749.full.pdf

Cloning of genomic and complementary DNA from Shaker, a putative potassium channel gene from Drosophila.

The hippocampal pyramidal cells provide an example of how multiple potassium (K) currents co-exist and function in central mammalian neurones. The data come from CA1 and CA3 neurones in hippocampal slices, cell cultures and acutely dissociated cells from rats and guinea-pigs. Six voltage- or calcium(Ca)-dependent K currents have so far been described in CA1 pyramidal cells in slices. Four of them (IA, ID, IK, IM) are activated by depolarization alone; the two others (IC, IAHP) are activated by voltage-dependent influx of Ca ions (IC may be both Ca- and voltage-gated). In addition, a transient Ca-dependent K current (ICT) has been described in certain preparations, but it is not yet clear whether it is distinct from IC and IA. (1) IA activates fast (within 10 ms) and inactivates rapidly (time constant typically 15-50 ms) at potentials positive to -60 mV; it probably contributes to early spike-repolarization, it can delay the first spike for about 0.1 s, and may regulate repetitive firing. (2) ID activates within about 20 ms but inactivates slowly (seconds) below the spike threshold (-90 to -60 mV), causing a long delay (0.5-5 s) in the onset of firing. Due to its slow recovery from inactivation (seconds), separate depolarizing inputs can be "integrated". ID probably also participates in spike repolarization. (3) IK activates slowly (time constant, tau, 20-60 ms) in response to depolarizations positive to -40 mV and inactivates (tau about 5s) at -80 to -40 mV; it probably participates in spike repolarization. (4) IM activates slowly (tau about 50 ms) positive to -60 mV and does not inactivate; it tends to attenuate excitatory inputs, it reduces the firing rate during maintained depolarization (adaptation) and contributes to the medium after-hyperpolarization (mAHP); IM is suppressed by acetylcholine (via muscarinic receptors), but may be enhanced by somatostatin. (5) IC is activated by influx of Ca ions during the action potential and is thought to cause the final spike repolarization and the fast AHP (although ICT may be involved). Like IM, it also contributes to the medium AHP and early adaptation. It differs from IAHP by being sensitive to tetraethylammonium (TEA, 1 mM), but insensitive to noradrenaline and muscarine. Large-conductance (BK; about 200 pS) Ca-activated K channels, which may mediate IC, have been recorded. (6) IAHP is slowly activated by Ca-influx during action potentials, causing spike-frequency adaptation and the slow AHP. Thus, IAHP exerts a strong negative feedback control of discharge activity.(ABSTRACT TRUNCATED AT 400 WORDS)

Potassium currents in hippocampal pyramidal cells.

1. The repolarization of the action potential, and a fast after-hyperpolarization (a.h.p.) were studied in CA1 pyramidal cells (n = 76) in rat hippocampal slices (28-37 degrees C). Single spikes were elicited by brief (1-3 ms) current pulses, at membrane potentials close to rest (-60 to -70 mV). 2. Each action potential was followed by four after-potentials: (a) the fast a.h.p., lasting 2-5 ms; (b) an after-depolarization; (c) a medium a.h.p., (50-100 ms); and (d) a slow a.h.p. (1-2 s). Both the fast a.h.p. and the slow a.h.p. (but not the medium a.h.p.) were inhibited by Ca2+-free medium or Ca2+-channel blockers (Co2+, Mn2+ or Cd2+); but tetraethylammonium (TEA; 0.5-2 nM) blocked only the fast a.h.p., and noradrenaline (2-5 microM) only the slow a.h.p. This suggests that two Ca2+-activated K+ currents were involved: a fast, TEA-sensitive one (IC) underlying the fast a.h.p., and a slow noradrenaline-sensitive one (IAHP) underlying the slow a.h.p. 3. Like the fast a.h.p., spike repolarization seems to depend on a Ca2+-dependent K+ current of the fast, TEA-sensitive kind (IC). The repolarization was slowed by Ca2+-free medium, Co2+, Mn2+, Cd2+, or TEA, but not by noradrenaline. Charybdotoxin (CTX; 30 nM), a scorpion toxin which blocks the large-conductance Ca2+-activated K+ channel in muscle, had a similar effect to TEA. The effects of TEA and Cd2+ (or Mn2+) showed mutual occlusion. Raising the external K+ concentration reduced the fast a.h.p. and slowed the spike repolarization, whereas Cl- loading of the cell was ineffective. 4. The transient K+ current, IA, seems also to contribute to spike repolarization, because: (a) 4-aminopyridine (4-AP; 0.1 mM), which blocks IA, slowed the spike repolarization; (b) depolarizing pre-pulses, which inactivate IA, had a similar effect; (c) hyperpolarizing pre-pulses speeded up the spike repolarization; (d) the effects of 4-AP and pre-pulses persisted during Ca2+ blockade (like IA); and (e) depolarizing pre-pulses reduced the effect of 4-AP. 5. Pre-pulses or 4-AP broadened the spike less, and in a different manner, than Ca2+-free medium, Cd2+, Co2+, Mn2+, TEA or CTX. The former broadening was uniform, with little effect on the fast a.h.p., whereas the latter affected mostly the last two-thirds of the spike repolarization and abolished the fast a.h.p.(ABSTRACT TRUNCATED AT 400 WORDS)

Action potential repolarization and a fast after‐hyperpolarization in rat hippocampal pyramidal cells.

Dendrotoxin, a small single-chain protein from the venom of Dendroaspis angusticeps, is highly toxic following intracerebroventricular injection into rats. Voltage-clamp analysis of CA1 neurons in hippocampal slices, treated with tetrodotoxin, revealed that nanomolar concentrations of dendrotoxin reduce selectively a transient, voltage-dependent K conductance. Epileptiform activity known to be induced by dendrotoxin can be attributed to such an action. Membrane currents not affected directly by the toxin include (i) Ca-activated K conductance; (ii) noninactivating voltage-dependent K conductance; (iii) inactivating and noninactivating Ca conductances; (iv) persistent inward (anomalous) rectifier current. Persistence of the effects of the toxin when Cd was included to suppress spontaneous transmitter release indicates a direct action on the neuronal membrane. Using biologically active, 125I-labeled dendrotoxin, protein acceptor sites of high affinity were detected on cerebrocortical synaptosomal membranes and sections of rat brain. In hippocampus, toxin binding was shown autoradiographically to reside in synapse-rich and white matter regions, with lower levels in cell body layers. This acceptor is implicated in the action of toxin because its affinities for dendrotoxin congeners are proportional to their central neurotoxicities and potencies in reducing the transient, voltage-dependent K conductance.

https://www.pnas.org/content/pnas/83/2/493.full.pdf

Central action of dendrotoxin: selective reduction of a transient K conductance in hippocampus and binding to localized acceptors

Homogeneous beta-bungarotoxin interacts irreversibly with rat olfactory cortex and produced permanent inhibition of neurotransmission (half-time of blockade for 230 nM toxin in 25 min). Binding occurs in the absence of divalent cations, but the rate of synaptic blockade is increased by Ca2+, which activates the intrinsic phospholipase A2 activity of the toxin. Other observable actions of the toxin, seen with rat cerebrocortical synaptosomes, are an increase in the release of acetylcholine, glutamate and gamma-aminobutyrate and impairment of transmitter uptake, which are all insensitive to tetrodotoxin. Inactivation of the toxin's phospholipase activity by chemical modification with p-bromophenacyl bromide diminishes the observed concomitant efflux of the neurotransmitters and lactate dehydrogenase. Collectively, the results support the idea that the toxin binds specifically and irreversibly to component(s) on nerve terminals and this together with the resultant phospholipolysis leads eventually to synaptic blockade. Such a proposal would account for the unique toxicity of the protein relative to phospholipase A2 enzymes.

Biochemical and Electrophysiological Demonstrations of the Actions of β‐Bungarotoxin on Synapses in Brain

β-Bungarotoxin, which was purified to homogeneity from the venom of Bungarus multicinctus by ion-exchange chromatography and isoelectric focussing, was shown to affect release of acetylcholine, glutamate and γ-aminobutyrate from rat cortex synaptosomes. In slices of rat olfactory cortex, the toxin irreversibly blocked neurotransmission by inhibiting transmitter release; in addition, it reduced the amplitude of the presynaptic action potential but at a slower rate. Inhibition of the toxin's phospholipase A2 activity, by replacement of Ca2+ with Sr2+, only attenuated its action on release of transmitters but abolished effects on the action potential. Irreversible binding of β-bungarotoxin occurred in Sr2+ and subsequent addition of Ca2+ restored its full neurotoxicity. Two pure phospholipases A2 were much less potent than toxin in mediating effects on transmitter release from the synaptosomes and slices. It is concluded that under certain conditions β-bungarotoxin can specifically inhibit transmitter release at a central synapse and appears, therefore, to be a useful probe for studying this process.

J. V. Halliwell

Papers

Central action of dendrotoxin: selective reduction of a transient K conductance in hippocampus and binding to localized acceptors

Biochemical and Electrophysiological Demonstrations of the Actions of β‐Bungarotoxin on Synapses in Brain

INTERACTION OF β-BUNGAROTOXIN WITH SYNAPSES IN THE MAMMALIAN CENTRAL NERVOUS SYSTEM