Daniel V. Madison

Multiple types of neuronal calcium channels and their selective modulation

Abstract Voltage-gated Ca 2+ channels control the entry of Ca 2+ ions across the surface membrane of neurons and other cells and thereby influence electrical activity and diverse cellular responses. Here we summarize efforts at classifying multiple types of Ca 2+ channels according to differences in their gating, ionic conductance and pharmacology. The distribution of Ca 2+ channel types among different cells and their selective modulation by neurochemicals and drugs is briefly reviewed.

Voltage clamp analysis of cholinergic action in the hippocampus

A slow muscarinic EPSP, accompanied by an increase in membrane input resistance, can be elicited in hippocampal CA1 pyramidal cells in vitro by electrical stimulation of cholinergic afferents in the slice preparation. Associated with the slow EPSP is a blockade of calcium- activated potassium afterhyperpolarizations (AHPs) (Cole and Nicoll, 1984a). In this study a single-electrode voltage clamp was used to examine the currents affected by activation of muscarinic receptors, using either bath application of carbachol or electrical stimulation of the cholinergic afferents. The 3 main findings of this study are that (1) of the 2 calcium-activated potassium currents (termed IAHP and IC) in hippocampal pyramidal cells, only IAHP is sensitive to carbachol; (2) IAHP is approximately 10-fold more sensitive to carbachol than is another muscarine-sensitive current, IM; and (3) neither blockade of IAHP nor of IM can account for the production of the slow EPSP. Rather, the slow EPSP appears to be generated by the blockade of a nonvoltage- dependent, resting potassium current. We propose that the muscarinic blockade of IAHP, which largely accounts for spike frequency adaptation, is primarily involved in enhancing action potential discharge to depolarizing stimuli, while the slow EPSP acts directly to cause action potential discharge.

Actions of noradrenaline recorded intracellularly in rat hippocampal CA1 pyramidal neurones, in vitro.

CA1 pyramidal neurones were studied in rat in vitro hippocampal slices using standard intracellular and single‐electrode voltage‐clamp recording techniques to examine the actions of noradrenaline (NA). NA had two different effects on the resting membrane potential of pyramidal neurones; either a hyperpolarization accompanied by a decrease in membrane input resistance, or less commonly, a depolarization accompanied by an increase in input resistance. In many cells, both effects, a hyperpolarization followed by a depolarization were observed. The depolarization was mediated by a noradrenergic beta‐receptor. The hyperpolarization was more difficult to characterize, but may result from alpha‐receptor activation. NA reduced the amplitude and duration of the slow calcium‐activated potassium after‐hyperpolarization (a.h.p.) that follows depolarization‐induced action potentials. This action of NA was mediated by beta 1‐noradrenergic receptors. NA, in the presence of tetrodotoxin and tetraethylammonium, reduced the a.h.p. without reducing the size of the calcium action potential which preceded it. This was unlike the action of the calcium channel blocker, cadmium, which reduced the calcium action potential and the a.h.p. in parallel. Furthermore, NA did not reduce the amplitude of calcium or barium currents recorded under voltage clamp after blockade of potassium currents. A functional consequence of this blockade of the calcium‐activated a.h.p. was a reduction of the accommodation of action potential discharge such that the excitatory responses of the neurone to depolarizing stimuli, such as glutamate application or current passed through the recording electrode, were enhanced. We conclude that the effects of NA on calcium‐activated potassium conductance and on resting membrane potential can interact to increase the signal‐to‐noise ratio of hippocampal pyramidal neurone responsiveness.

Imaging of cytosolic Ca2+ transients arising from Ca2+ stores and Ca2+ channels in sympathetic neurons.

Changes in cytosolic free Ca2+ concentration [( Ca2+]i) due to Ca2+ entry or Ca2+ release from internal stores were spatially resolved by digital imaging with the Ca2+ indicator fura-2 in frog sympathetic neurons. Electrical stimulation evoked a rise in [Ca2+]i spreading radially from the periphery to the center of the soma. Elevated [K+]o also increased [Ca2+]i, but only in the presence of external Ca2+, indicating that Ca2+ influx through Ca2+ channels is the primary event in the depolarization response. Ca2+ release or uptake from caffeine-sensitive internal stores was able to amplify or attenuate the effects of Ca2+ influx, to generate continued oscillations in [Ca2+]i, and to persistently elevate [Ca2+]i above basal levels after the stores had been Ca2(+)-loaded.

Enkephalin hyperpolarizes interneurones in the rat hippocampus.

1. Intracellular recordings were made from pyramidal cells and from electrophysiologically identified interneurones in the CA1 region of the hippocampal slice preparation from the rat. 2. Enkephalin blocked the hyperpolarization of pyramidal cells evoked by application of glutamate to synaptically coupled inhibitory interneurones. 3. Enkephalin hyperpolarized interneurones, most probably by increasing potassium conductance; this action was blocked by the opiate antagonist, naloxone. 4. Activation of gamma‐aminobutyric acid(B) receptors with baclofen in interneurones produced a similar hyperpolarization that was resistant to naloxone. 5. In addition to hyperpolarizing interneurones, enkephalin blocked the inhibitory postsynaptic potential recorded in these cells. 6. These results suggest that opiate receptors are selectively localized on inhibitory interneurones in the hippocampus and are coupled to potassium channels. Activation of these receptors causes a disinhibition of both pyramidal cells and inhibitory interneurones.

Nicotinic Receptor Activation Excites Distinct Subtypes of Interneurons in the Rat Hippocampus

We examined the function of nicotinic acetylcholine receptors (nAChRs) in interneurons of area CA1 of the rat hippocampus. CA1 interneurons could be classified into three categories based on nicotinic responses. The first class was depolarized by α7 nAChRs, found in all layers of CA1 and as a group, had axonal projections to all neuropil layers of CA1. The second class had both fast α7 and slow non-α7 nAChR depolarizing responses, was localized primarily to the stratum oriens, and had axonal projections to the stratum lacunosum-moleculare. The third group had no nicotinic response. This group was found in or near the stratum pyramidale and had axonal projections almost exclusively within and around this layer. Low concentrations (500 nm) of nicotine desensitized fast and slow nAChR responses. These findings demonstrate that there are distinct subsets of interneurons with regard to nicotinic receptor expression and with predictable morphological properties that suggest potential cellular actions for nicotinic receptor activation in normal CNS function and during nicotine abuse.

Opioid inhibition of GABA release from presynaptic terminals of rat hippocampal interneurons.

Opiates and the opioid peptide enkephalin can cause indirect excitation of principal cortical neurons by reducing inhibitory synaptic transmission mediated by GABAergic interneurons. The mechanism by which opioids mediate these effects on interneurons is unknown, but enkephalin hyperpolarizes the somatic membrane potential of a variety of neurons in the brain, including hippocampal interneurons. We now report a new, more direct mechanism for the opioid-mediated reduction in synaptic inhibition. The enkephalin analog D-Ala2-Met5-enkephalinamide (DALA) decreases the frequency of miniature, action potential-independent, spontaneous GABAergic inhibitory postsynaptic currents (IPSCs) without causing a change in their amplitude. Thus, we conclude that DALA inhibits the action potential-independent release of GABA through a direct action on interneuronal synaptic terminals. In contrast, DALA reduces the amplitude of action potential-evoked, GABA-mediated IPSCs, as well as decreases their frequency. This suggests that the opioid-mediated inhibition of non-action potential-dependent GABA release reveals a mechanism that contributes to reducing action potential-evoked GABA release, thereby decreasing synaptic inhibition.

Inhibition of hippocampal heme oxygenase, nitric oxide synthase, and long-term potentiation by metalloporphyrins

Four potent metalloporphyrin inhibitors of heme oxygenase were used to assess whether carbon monoxide production was required for induction of LTP in the CA1 region of the hippocampus. Although the metalloporphyrins produced a similar and substantial inhibition of heme oxygenase activity in hippocampal slices, only two compounds reduced the amount of LTP elicited by tetanic stimulation (chromium mesoporphyrin IX and zinc protoporphyrin IX). Both chromium mesoporphyrin IX and zinc protoporphyrin IX inhibited nitric oxide synthase in the hippocampus; tin mesoporphyrin IX and zinc deuteroporphyrin IX bis glycol neither reduced LTP induction nor inhibited NOS activity, although they did inhibit heme oxygenase. None of these metalloporphyrins reversed established LTP. Thus, together these data do not support carbon monoxide as a mediator in either LTP induction or expression/maintenance and emphasize further the nonselectivity of some metalloporphyrins.

Pair recordings reveal all-silent synaptic connections and the postsynaptic expression of long-term potentiation.

The activation of silent synapses is a proposed mechanism to account for rapid increases in synaptic efficacy such as long-term potentiation (LTP). Using simultaneous recordings from individual pre- and postsynaptic neurons in organotypic hippocampal slices, we show that two CA3 neurons can be connected entirely by silent synapses. Increasing release probability or application of cyclothiazide does not produce responses from these silent synapses. Direct measurement of NMDAR-mediated postsynaptic responses in all-silent synaptic connections before and after LTP induction show no change in failure rate, amplitude, or area. These data do not support hypotheses that synapse silent results from presynaptic factors or that LTP results from increases in presynaptic glutamate release. LTP is also associated with an increase in postsynaptic responsiveness to exogenous AMPA. We conclude that synapse silence, activation, and expression of LTP are postsynaptic.

Reflections on Ca2+-channel diversity, 1988–1994

partners throughout the CNS. Added to this, the coupling of the NO-cGMP pathway to ectlvatlon of glutamate [especially N-methyl-n-aspartate (NMDA)] receptors, first demonstrated In the cerebellum*, appears to be widely applicable. Flnally, the early avallablllty of pharmacologlcal tools, notably lnhlbltors of NO synthase, NO scav. engers and drugs that donate NO, provlded the means to test for the lnvolvement of NO In CNS function. It turns out to be lmpllcated In a bewIlder. lng array of phenomena, ranging from synaptic plastlclty In several of Its forms (for example, long4erm potentlatlon and depresslon of syneptlc transmls= slon, synaptogenesls and development, drug tolerance and dependence and hyperalgesla) to the moduIatlon of sensory nnd motor pathways, the local regulation of cerebral blood flow, neuroendocrlne reguiatlon, leernlng, feedlng and sexual behavlour’“, Under more pathologlcal condltlons, excessive productlon of NO by neurones or gllrl cells might be an lmportnnt medlator of cell denth nnoI thur, contrlbute to ncurodcgeneratlve states, for cnar~ple, followlng cerebral lschaemla~4~~~, The field Is relatively young, and It Is still expand. Ing rapldly: the past few years have seen an annual twoto threefold Increase In the number of papers publlshed on the subject, Dssplte the Increase In knowledge, our level of understandlng of NO slgnalllng In the braln remains crude, Many key Issues surroundlng Its proposed functions remeln unresolved and steeped In controversy, for example, Its role (or otherwlse) In Ion&term potentlatlon and Its supposed whole-animal correlates of leemlng and memory. The dust should settle, In tlme. One of the most Important goals for the future Is to elucidate the cellular end molecular mechenlsms performed by NO end Its second messenger, cGMP, In neural cells, Wlth respect to the latter, and wlth a few notable exceptlons, we are not much wiser than we were when It was first found In the braln, 25 years ago,