Slow Afterhyperpolarization Current Research Articles

Vasoactive intestinal peptide excites GnRH neurons via KCa3.1, a potential player in the slow afterhyperpolarization current.

Vasoactive intestinal peptide (VIP) is an important component of the suprachiasmatic nucleus (SCN) which relays circadian information to neuronal populations, including GnRH neurons. Human and animal studies have shown an impact of disrupted daily rhythms (chronic shift work, temporal food restriction, clock gene disruption) on both male and female reproduction and fertility. To date, how VIP modulates GnRH neurons remains unknown. Calcium imaging and electrophysiology on primary GnRH neurons in explants and adult mouse brain slice, respectively, were used to address this question. We found VIP excites GnRH neurons via the VIP receptor, VPAC2. The downstream signaling pathway uses both Gs protein/adenylyl cyclase/protein kinase A (PKA) and phospholipase C/phosphatidylinositol 4,5-bisphosphate (PIP2) depletion. Furthermore, we identified a UCL2077-sensitive target, likely contributing to the slow afterhyperpolarization current (IAHP), as the PKA and PIP2 depletion target, and the KCa3.1 channel as a specific target. Thus, VIP/VPAC2 provides an example of Gs protein-coupled receptor-triggered excitation in GnRH neurons, modulating GnRH neurons likely via the slow IAHP. The possible identification of KCa3.1 in the GnRH neuron slow IAHP may provide a new therapeutical target for fertility treatments.

Altered calcium signaling in Bergmann glia contributes to spinocerebellar ataxia type-1 in a mouse model of SCA1

Spinocerebellar ataxia type 1 (SCA1) is a neurodegenerative disease caused by an abnormal expansion of glutamine (Q) encoding CAG repeats in the ATAXIN1 (ATXN1) gene and characterized by progressive cerebellar ataxia, dysarthria, and eventual deterioration of bulbar functions. SCA1 shows severe degeneration of cerebellar Purkinje cells (PCs) and activation of Bergmann glia (BG), a type of cerebellar astroglia closely associated with PCs. Combining electrophysiological recordings, calcium imaging techniques, and chemogenetic approaches, we have investigated the electrical intrinsic and synaptic properties of PCs and the physiological properties of BG in SCA1 mouse model expressing mutant ATXN1 only in PCs. PCs of SCA1 mice displayed lower spontaneous firing rate and larger slow afterhyperpolarization currents (sIAHP) than wildtype mice, whereas the properties of the synaptic inputs were unaffected. BG of SCA1 mice showed higher calcium hyperactivity and gliotransmission, manifested by higher frequency of NMDAR-mediated slow inward currents (SICs) in PC. Preventing the BG calcium hyperexcitability of SCA1 mice by loading BG with the calcium chelator BAPTA restored sIAHP and spontaneous firing rate of PCs to similar levels of wildtype mice. Moreover, mimicking the BG hyperactivity by activating BG expressing Gq-DREADDs in wildtype mice reproduced the SCA1 pathological phenotype of PCs, i.e., enhancement of sIAHP and decrease of spontaneous firing rate. These results indicate that the intrinsic electrical properties of PCs, but not their synaptic properties, were altered in SCA1 mice and that these alterations were associated with the hyperexcitability of BG. Moreover, preventing BG hyperexcitability in SCA1 mice and promoting BG hyperexcitability in wildtype mice prevented and mimicked, respectively, the pathological electrophysiological phenotype of PCs. Therefore, BG plays a relevant role in the dysfunction of the electrical intrinsic properties of PCs in SCA1 mice, suggesting that they may serve as potential targets for therapeutic approaches to treat the spinocerebellar ataxia type 1.

GluK2 Q/R editing regulates kainate receptor signaling and long-term potentiation of AMPA receptors

Q/R editing of the kainate receptor (KAR) subunit GluK2 radically alters recombinant KAR properties, but the effects on endogenous KARs invivo remain largely unexplored. Here, we compared GluK2 editing-deficient mice that express ∼95% unedited GluK2(Q) to wild-type counterparts that express ∼85% edited GluK2(R). At mossy fiber-CA3 (MF-CA3) synapses GluK2(Q) mice displayed increased postsynaptic KAR function and KAR-mediated presynaptic facilitation, demonstrating enhanced ionotropic function. Conversely, GluK2(Q) mice exhibited reduced metabotropic KAR function, assessed by KAR-mediated inhibition of slow after-hyperpolarization currents (ISAHP). GluK2(Q) mice also had fewer GluA1-and GluA3-containing AMPA receptors (AMPARs) and reduced postsynaptic AMPAR currents at both MF-CA3 and CA1-Schaffer collateral synapses. Moreover, long-term potentiation of AMPAR-mediated transmission at CA1-Schaffer collateral synapses was reduced in GluK2(Q) mice. These findings suggest that GluK2 Q/R editing influences ionotropic/metabotropic balance of KAR signaling to regulate synaptic expression of AMPARs and plasticity.

Noradrenaline Release from Locus Coeruleus Terminals in the Hippocampus Enhances Excitation-Spike Coupling in CA1 Pyramidal Neurons Via β-Adrenoceptors.

Release of the neuromodulator noradrenaline signals salience during wakefulness, flagging novel or important experiences to reconfigure information processing and memory representations in the hippocampus. Noradrenaline is therefore expected to enhance hippocampal responses to synaptic input; however, noradrenergic agonists have been found to have mixed and sometimes contradictory effects on Schaffer collateral synapses and the resulting CA1 output. Here, we examine the effects of endogenous, optogenetically driven noradrenaline release on synaptic transmission and spike output in mouse hippocampal CA1 pyramidal neurons. We show that endogenous noradrenaline release enhances the probability of CA1 pyramidal neuron spiking without altering feedforward excitatory or inhibitory synaptic inputs in the Schaffer collateral pathway. β-adrenoceptors mediate this enhancement of excitation-spike coupling by reducing the charge required to initiate action potentials, consistent with noradrenergic modulation of voltage-gated potassium channels. Furthermore, we find the likely effective concentration of endogenously released noradrenaline is sub-micromolar. Surprisingly, although comparable concentrations of exogenous noradrenaline cause robust depression of slow afterhyperpolarization currents, endogenous release of noradrenaline does not, indicating that endogenous noradrenaline release is targeted to specific cellular locations. These findings provide a mechanism by which targeted endogenous release of noradrenaline can enhance information transfer in the hippocampus in response to salient events.

Sodium and potassium conductances in principal neurons of the mouse piriform cortex: a quantitative description.

The primary olfactory (or piriform) cortex is a promising model system for understanding how the cerebral cortex processes sensory information, although an investigation of the piriform cortex is hindered by a lack of detailed information about the intrinsic electrical properties of its component neurons. In the present study, we quantify the properties of voltage-dependent sodium currents and voltage- and calcium-dependent potassium currents in two important classes of excitatory neurons in the main input layer of the piriform cortex. We identify several classes of these currents and show that their properties are similar to those found in better-studied cortical regions. Our detailed quantitative descriptions of these currents will be valuable to computational neuroscientists who aim to build models that explain how the piriform cortex encodes odours. The primary olfactory cortex (or piriform cortex, PC) is an anatomically simple palaeocortex that is increasingly used as a model system for investigating cortical sensory processing. However, little information is available on the intrinsic electrical conductances in neurons of the PC, hampering efforts to build realistic computational models of this cortex. In the present study, we used nucleated macropatches and whole-cell recordings to rigorously quantify the biophysical properties of voltage-gated sodium (NaV ), voltage-gated potassium (KV ) and calcium-activated potassium (KCa ) conductances in two major classes of glutamatergic neurons in layer 2 of the PC, semilunar (SL) cells and superficial pyramidal (SP) cells. We found that SL and SP cells both express a fast-inactivating NaV current, two types of KV current (A-type and delayed rectifier-type) and three types of KCa current (fast-, medium- and slow-afterhyperpolarization currents). The kinetic and voltage-dependent properties of the NaV and KV conductances were, with some exceptions, identical in SL and SP cells and similar to those found in neocortical pyramidal neurons. The KCa conductances were also similar across the different types of neurons. Our results are summarized in a series of empirical equations that should prove useful to computational neuroscientists seeking to model the PC. More broadly, our findings indicate that, at the level of single-cell electrical properties, this palaeocortex is not so different from the neocortex, vindicating efforts to use the PC as a model of cortical sensory processing in general.

Kisspeptin inhibits a slow afterhyperpolarization current via protein kinase C and reduces spike frequency adaptation in GnRH neurons

Kisspeptin signaling via its cognate receptor G protein-coupled receptor 54 (GPR54) in gonadotropin-releasing hormone (GnRH) neurons plays a critical role in regulating pituitary secretion of luteinizing hormone and thus reproductive function. GPR54 is G(q)-coupled to activation of phospholipase C and multiple second messenger signaling pathways. Previous studies have shown that kisspeptin potently depolarizes GnRH neurons through the activation of canonical transient receptor potential channels and inhibition of inwardly rectifying K(+) channels to generate sustained firing. Since the initial studies showing that kisspeptin has prolonged effects, the question has been why is there very little spike frequency adaption during sustained firing? Presently, we have discovered that kisspeptin reduces spike frequency adaptation and prolongs firing via the inhibition of a calcium-activated slow afterhyperpolarization current (I(sAHP)). GnRH neurons expressed two distinct I(sAHP), a kisspeptin-sensitive and an apamin-sensitive I(sAHP). Essentially, kisspeptin inhibited 50% of the I(sAHP) and apamin inhibited the other 50% of the current. Furthermore, the kisspeptin-mediated inhibition of I(sAHP) was abrogated by the protein kinase C (PKC) inhibitor calphostin C, and the PKC activator phorbol 12,13-dibutyrate mimicked and occluded any further effects of kisspeptin on I(sAHP). The protein kinase A (PKA) inhibitors H-89 and the Rp diastereomer of adenosine 3',5'-cyclic monophosphorothioate had no effect on the kisspeptin-mediated inhibition but were able to abrogate the inhibitory effects of forskolin on the I(sAHP), suggesting that PKA is not involved. Therefore, in addition to increasing the firing rate through an overt depolarization, kisspeptin can also facilitate sustained firing through inhibiting an apamin-insensitive I(sAHP) in GnRH neurons via a PKC.

Network properties of a computational model of the dorsal raphe nucleus

Serotonin (5-HT) plays an important role in regulating mood, cognition and behaviour. The midbrain dorsal raphe nucleus (DRN) is one of the primary sources of 5-HT. Recent studies show that DRN neuronal activities can encode rewarding (e.g., appetitive) and unrewarding (e.g., aversive) behaviours. Experiments have also shown that DRN neurons can exhibit heterogeneous spiking behaviours. In this work, we build and study a basic spiking neuronal network model of the DRN constrained by neuronal properties observed in experiments. We use an efficient adaptive quadratic integrate-and-fire neuronal model to capture slow afterhyperpolarization current, occasional bursting behaviours in 5-HT neurons, and fast spiking activities in the non-5-HT inhibitory neurons. Provided that our noisy and heterogeneous spiking neuronal network model adopts a feedforward inhibitory network architecture, it is able to replicate the main features of DRN neuronal activities recorded in monkeys performing a reward-based memory-guided saccade task. The model exhibits theta band oscillation, especially among the non-5-HT inhibitory neurons during the rewarding outcome of a simulated trial, thus forming a model prediction. By varying the inhibitory synaptic strengths and the afferent inputs, we find that the network model can oscillate over a range of relatively low frequencies, allow co-existence of multiple stable frequencies, and spike synchrony can spread from within a local neural subgroup to global. Our model suggests plausible network architecture, provides interesting model predictions that can be experimentally tested, and offers a sufficiently realistic multi-scale model for 5-HT neuromodulation simulations.

Glutamate transporters control metabotropic glutamate receptors activation to prevent the genesis of paroxysmal burst in the developing hippocampus

Metabotropic glutamate receptors (mGluR) can control neuronal excitability by modulating several ionic channels. In hippocampal pyramidal cells, groups I/II mGluR are located extrasynaptically, suggesting that their endogenous activation is dependent on the glutamate clearance rate and therefore on excitatory amino-acid transporters (EAAT) efficiency. Deficiency of glutamate uptake can generate seizures in rodents and has been suggested as a mechanism of seizure generation in some human epileptic syndromes. However, the cellular mechanisms linking EAAT dysfunction and pathological cortical activities remain elusive. Here, we investigate the possible role of mGluR on paroxysmal burst of multiple unit activities (MUA) generated in the CA1 region of developing hippocampal slices using an EAAT inhibitor, TBOA. These bursts are generated by a synaptic release of glutamate and involve extrasynaptic NMDA receptors (NMDAR) activated by transmitter spillover. Here, we show that postsynaptic mGluR (groups I/II) are tonically activated by the rise in ambient glutamate concentration after EAAT inhibition and strongly contribute to paroxysmal burst genesis. The inhibition of mGluR with broad spectrum antagonists or addition of a glutamate scavenger strongly reduced the occurrence of paroxysmal burst and the frequency/number of MUA during the burst. Moreover, this endogenous activation of groups I/II mGluR leads to (i) the reduction of the slow afterhyperpolarization current (IsAHP), increasing the firing pattern of pyramidal cells, and (ii) the potentiation of extrasynaptic NMDAR-mediated responses, enabling glutamate spillover to generate a suprathreshold depolarization for several seconds. Our data show that an insufficient buffering of extracellular glutamate enables a cross talk between groups I/II mGluR and NMDAR, which, combined with a decrease of IsAHP, leads to the hyperexcitability of the hippocampal network, facilitating the genesis of epileptical-like activity in response to glutamate release. These findings highlight the importance of the control exerted by EAAT on mGluR.

Persistence and storage of activity patterns in spiking recurrent cortical networks: modulation of sigmoid signals by after-hyperpolarization currents and acetylcholine.

Many cortical networks contain recurrent architectures that transform input patterns before storing them in short-term memory (STM). Theorems in the 1970's showed how feedback signal functions in rate-based recurrent on-center off-surround networks control this process. A sigmoid signal function induces a quenching threshold below which inputs are suppressed as noise and above which they are contrast-enhanced before pattern storage. This article describes how changes in feedback signaling, neuromodulation, and recurrent connectivity may alter pattern processing in recurrent on-center off-surround networks of spiking neurons. In spiking neurons, fast, medium, and slow after-hyperpolarization (AHP) currents control sigmoid signal threshold and slope. Modulation of AHP currents by acetylcholine (ACh) can change sigmoid shape and, with it, network dynamics. For example, decreasing signal function threshold and increasing slope can lengthen the persistence of a partially contrast-enhanced pattern, increase the number of active cells stored in STM, or, if connectivity is distance-dependent, cause cell activities to cluster. These results clarify how cholinergic modulation by the basal forebrain may alter the vigilance of category learning circuits, and thus their sensitivity to predictive mismatches, thereby controlling whether learned categories code concrete or abstract features, as predicted by Adaptive Resonance Theory. The analysis includes global, distance-dependent, and interneuron-mediated circuits. With an appropriate degree of recurrent excitation and inhibition, spiking networks maintain a partially contrast-enhanced pattern for 800 ms or longer after stimuli offset, then resolve to no stored pattern, or to winner-take-all (WTA) stored patterns with one or multiple winners. Strengthening inhibition prolongs a partially contrast-enhanced pattern by slowing the transition to stability, while strengthening excitation causes more winners when the network stabilizes.

After-hyperpolarization currents and acetylcholine control sigmoid transfer functions in a spiking cortical model

Recurrent networks are ubiquitous in the brain, where they enable a diverse set of transformations during perception, cognition, emotion, and action. It has been known since the 1970's how, in rate-based recurrent on-center off-surround networks, the choice of feedback signal function can control the transformation of input patterns into activity patterns that are stored in short term memory. A sigmoid signal function may, in particular, control a quenching threshold below which inputs are suppressed as noise and above which they may be contrast enhanced before the resulting activity pattern is stored. The threshold and slope of the sigmoid signal function determine the degree of noise suppression and of contrast enhancement. This article analyses how sigmoid signal functions and their shape may be determined in biophysically realistic spiking neurons. Combinations of fast, medium, and slow after-hyperpolarization (AHP) currents, and their modulation by acetylcholine (ACh), can control sigmoid signal threshold and slope. Instead of a simple gain in excitability that was previously attributed to ACh, cholinergic modulation may cause translation of the sigmoid threshold. This property clarifies how activation of ACh by basal forebrain circuits, notably the nucleus basalis of Meynert, may alter the vigilance of category learning circuits, and thus their sensitivity to predictive mismatches, thereby controlling whether learned categories code concrete or abstract information, as predicted by Adaptive Resonance Theory.

VIP, CRF, and PACAP Act at Distinct Receptors to Elicit Different cAMP/PKA Dynamics in the Neocortex

The functional significance of diverse neuropeptide coexpression and convergence onto common second messenger pathways remains unclear. To address this question, we characterized responses to corticotropin-releasing factor (CRF), pituitary adenylate cyclase-activating peptide (PACAP), and vasoactive intestinal peptide (VIP) in rat neocortical slices using optical recordings of cyclic adenosine monophosphate (cAMP) and protein kinase A (PKA) sensors, patch-clamp, and single-cell reverse transcription-polymerase chain reaction. Responses of pyramidal neurons to the 3 neuropeptides markedly differed in time-course and amplitude. Effects of these neuropeptides on the PKA-sensitive slow afterhyperpolarization current were consistent with those observed with cAMP/PKA sensors. CRF-1 receptors, primarily expressed in pyramidal cells, reportedly mediate the neocortical effects of CRF. PACAP and VIP activated distinct PAC1 and VPAC1 receptors, respectively. Indeed, a selective VPAC1 antagonist prevented VIP responses but had a minor effect on PACAP responses, which were mimicked by a specific PAC1 agonist. While PAC1 and VPAC1 were coexpressed in pyramidal cells, PAC1 expression was also frequently detected in interneurons, suggesting that PACAP has widespread effects on the neuronal network. Our results suggest that VIP and CRF, originating from interneurons, and PACAP, expressed mainly by pyramidal cells, finely tune the excitability and gene expression in the neocortical network via distinct cAMP/PKA-mediated effects.

The KCNQ5 potassium channel mediates a component of the afterhyperpolarization current in mouse hippocampus

Mutations in KCNQ2 and KCNQ3 voltage-gated potassium channels lead to neonatal epilepsy as a consequence of their key role in regulating neuronal excitability. Previous studies in the brain have focused primarily on these KCNQ family members, which contribute to M-currents and afterhyperpolarization conductances in multiple brain areas. In contrast, the function of KCNQ5 (Kv7.5), which also displays widespread expression in the brain, is entirely unknown. Here, we developed mice that carry a dominant negative mutation in the KCNQ5 pore to probe whether it has a similar function as other KCNQ channels. This mutation renders KCNQ5(dn)-containing homomeric and heteromeric channels nonfunctional. We find that Kcnq5(dn/dn) mice are viable and have normal brain morphology. Furthermore, expression and neuronal localization of KCNQ2 and KCNQ3 subunits are unchanged. However, in the CA3 area of hippocampus, a region that highly expresses KCNQ5 channels, the medium and slow afterhyperpolarization currents are significantly reduced. In contrast, neither current is affected in the CA1 area of the hippocampus, a region with low KCNQ5 expression. Our results demonstrate that KCNQ5 channels contribute to the afterhyperpolarization currents in hippocampus in a cell type-specific manner.

Two Slow Calcium-Activated Afterhyperpolarization Currents Control Burst Firing Dynamics in Gonadotropin-Releasing Hormone Neurons

Gonadotropin-releasing hormone (GnRH) neurons release GnRH in a pulsatile manner to control fertility in all mammals. The mechanisms underlying burst firing in GnRH neurons, thought to contribute to pulsatile GnRH release, are not yet understood. Using minimally invasive, dual electrical-calcium recordings in acute brain slices from GnRH-Pericam transgenic mice, we find that the soma/proximal dendrites of GnRH neurons exhibit long-duration (approximately 10 s) calcium transients that are perfectly synchronized with their burst firing. These transients were found to be generated by calcium entry through voltage-dependent L-type calcium channels that was amplified by inositol-1,4,5-trisphosphate receptor-dependent store mechanisms. Perforated-patch current- and voltage-clamp electrophysiology coupled with mathematical modeling approaches revealed that these broad calcium transients act to control two slow afterhyperpolarization currents (sI(AHP)) in GnRH neurons: a quick-activating apamin-sensitive sI(AHP) that regulates both intraburst and interburst dynamics, and a slow-onset UCL2077-sensitive sI(AHP) that regulates only interburst dynamics. These observations highlight a unique interplay between electrical activity, calcium dynamics, and multiple calcium-regulated sI(AHP)s critical for shaping GnRH neuron burst firing.

High-Affinity Kainate Receptor Subunits Are Necessary for Ionotropic but Not Metabotropic Signaling

Kainate receptors signal through both ionotropic and metabotropic pathways. The high-affinity subunits, GluK4 and GluK5, are unique among the five receptor subunits, as they do not form homomeric receptors but modify the properties of heteromeric assemblies. Disruption of the Grik4 gene locus resulted in a significant reduction in synaptic kainate receptor currents. Moreover, ablation of GluK4 and GluK5 caused complete loss of synaptic ionotropic kainate receptor function. The principal subunits were distributed away from postsynaptic densities and presynaptic active zones. There was also a profound alteration in the activation properties of the remaining kainate receptors. Despite this, kainate receptor-mediated inhibition of the slow afterhyperpolarization current (I(sAHP)), which is dependent on metabotropic pathways, was intact in GluK4/GluK5 knockout mice. These results uncover a previously unknown obligatory role for the high-affinity subunits for ionotropic kainate receptor function and further demonstrate that kainate receptor participation in metabotropic signaling pathways does not require their classic role as ion channels.

Contribution of KCNQ2 and KCNQ3 to the medium and slow afterhyperpolarization currents

Benign familial neonatal convulsion (BNFC) is a neurological disorder caused by mutations in the potassium channel genes KCNQ2 and KCNQ3, which are thought to contribute to the medium afterhyperpolarization (mAHP). Despite their importance in normal brain function, it is unknown whether they invariably function as heteromeric complexes. Here, we examined the contribution of KCNQ3 and KCNQ2 in mediating the apamin-insensitive mAHP current (ImAHP) in hippocampus. The ImAHP was not impaired in CA1 pyramidal neurons from mice genetically deficient for either KCNQ3 or KCNQ2 but was reduced approximately 50% in dentate granule cells. While recording from KCNQ-deficient mice, we observed that the calcium-activated slow afterhyperpolarization current (IsAHP) was also reduced in dentate granule cells, suggesting that KCNQ channels might also contribute to this potassium current whose molecular identity is unknown. Further pharmacological and molecular experiments manipulating KCNQ channels provided evidence in support of this possibility. Together our data suggest that multiple KCNQ subunit compositions can mediate the ImAHP, and that the very same subunits may also contribute to the IsAHP. We also present data suggesting that the neuronal calcium sensor protein hippocalcin may allow for these dual signaling processes.

Rat GnRH Neurons Exhibit Large Conductance Voltage- and Ca2+-Activated K+ (BK) Currents and Express BK Channel mRNAs

Gonadotropin-releasing hormone (GnRH) neurons form the final common pathway for the central regulation of reproduction. As in other neurons, the discharge pattern of action potentials is important for these neurons to function properly. Therefore it is important to elucidate the expression patterns of various types of ion channels in these neurons because they determine cell excitability. To date, voltage-gated Ca2+ channels and SK channels have been reported to be expressed in rat GnRH neurons. In this study, we focused on K+ channels and analyzed their expression in primary cultured GnRH neurons, prepared from GnRH-EGFP transgenic rats, by means of perforated patch-clamp recordings. GnRH neurons exhibited delayed-rectifier K+ currents and large conductance voltage- and Ca2+-activated K+ (BK) currents. Moreover, multicell RT-PCR (reverse transcriptase-polymerase chain reaction) experiments revealed the expression of BK channel mRNAs (alpha, beta1, beta2, and beta4). The results show the presence of delayed-rectifier K+ currents and BK currents besides previously reported slow afterhyperpolarization currents. These currents control the action potential repolarization and probably also the firing pattern, thereby regulating the cell excitability of GnRH neurons.

Control of IsAHP in mouse hippocampus CA1 pyramidal neurons by RyR3-mediated calcium-induced calcium release

In several neuronal preparations, the ryanodine-sensitive calcium store was reported to participate in the generation of slow afterhyperpolarization currents (IsAHP) involved in spike frequency adaptation. We show that calcium release from the ryanodine-sensitive calcium store is a major determinant of the triggering of IsAHP in mouse CA1 pyramidal neurons. Whole-cell patch clamp recordings in hippocampus slices show that the intracellular calcium stores depletion using an inhibitor of the endoplasmic reticulum Ca2+-ATPase (5 microM cyclopiazonic acid), as well as the specific blockade of ryanodine receptors (100 microM ryanodine) both reduced the IsAHP by about 70%. Immunohistology, using an anti-RyR3 specific antibody, indicates that RyR3 expression is particularly enriched in the CA1 apical dendrites (considered as the most important site for sAHP generation). We show that our anti-RyR3 antibody acts as a functional RyR3 antagonist and induced a reduction in IsAHP by about 70%. The additional ryanodine application (100 micro M) did not further affect IsAHP, thus excluding RyR2 in IsAHP activation. Our results argue in favor of a specialized function of RyR3 in CA1 pyramidal cells in triggering IsAHP due to their localization in the apical dendrite.

The Calcium Sensor for Slow Afterhyperpolarization

Neurons in the brain protect themselves from excessive activity in part by a mechanism through which an influx of calcium ions activates potassium channels, which then cause hyperpolarization of the nerve cell, thus inhibiting further action potentials. The identity of this potassium channel is not known, but the current it conducts is called the slow afterhyperpolarization current (I sAHP ). Tzingounis et al . identified a new key component of the mechanism by which calcium influences the channel: the neuronal calcium sensor family member known as hippocalcin. The authors interpreted results from kinetic analysis of I sAHP in the presence of various calcium buffers to indicate that I sAHP responded primarily to general changes in calcium concentration within the cell, as opposed to large rapid changes in calcium concentration in the immediate vicinity of calcium channels. The latter is characteristic of other potassium channels that exist together with calcium channels in large complexes in which the calcium sensor is bound to or part of the potassium channel. Instead, the sAHP channels appeared to be regulated by a diffusible calcium sensor, so the authors suspected hippocalcin. They found that, indeed, hippocampal slices from hippocalcin knockout mice lacked I sAHP , whereas expression of hippocalcin in cultured neurons enhanced I sAHP . Hippocalcin contains a "myristoyl switch," a mechanism by which a conformational change induced by calcium binding causes exposure of a myristoyl moiety that causes hippocalcin to bind to the plasma membrane. A mutant lacking the myristoylation site failed to increase I sAHP . The authors therefore propose that hippocalcin senses global changes in the concentration of intracellular free calcium and relays that information back to the plasma membrane. Brown et al . point out in commentary that elucidation of the mechanism by which hippocalcin actually regulates the sAHP channel, be it direct or through another messenger molecule (hippocalcin is known to bind phosphatidylinositol 4,5-bisphosphate), may be possible only when the so-far-elusive molecular identity of the sAHP channel itself is resolved. A. V. Tzingounis, M. Kobayashi, K. Takamatsu, R. A. Nicoll, Hippocalcin gates the calcium activation of the slow afterhyperpolarization in hippocampal pyramidal cells. Neuron 53 , 487-493 (2007). [Online Journal] D. A. Brown, B. Lancaster, M. M. Shah, Hippocalcin: A new solution to an old puzzle. Neuron 53 , 467-468 (2007). [Online Journal]

Hippocalcin Gates the Calcium Activation of the Slow Afterhyperpolarization in Hippocampal Pyramidal Cells

In the brain, calcium influx following a train of action potentials activates potassium channels that mediate a slow afterhyperpolarization current (I(sAHP)). The key steps between calcium influx and potassium channel activation remain unknown. Here we report that the key intermediate between calcium and the sAHP channels is the diffusible calcium sensor hippocalcin. Brief depolarizations sufficient to activate the I(sAHP) in wild-type mice do not elicit the I(sAHP) in hippocalcin knockout mice. Introduction of hippocalcin in cultured hippocampal neurons leads to a pronounced I(sAHP), while neurons expressing a hippocalcin mutant lacking N-terminal myristoylation exhibit a small I(sAHP) that is similar to that recorded in uninfected neurons. This implies that hippocalcin must bind to the plasma membrane to mediate its effects. These findings support a model in which the calcium sensor for the sAHP channels is not preassociated with the channel complex.

Stability and plasticity of intrinsic membrane properties in hippocampal CA1 pyramidal neurons: effects of internal anions

CA1 pyramidal neurons from animals that have acquired hippocampal tasks show increased neuronal excitability, as evidenced by a reduction in the postburst afterhyperpolarization (AHP). Studies of AHP plasticity require stable long-term recordings, which are affected by the intracellular solutions potassium methylsulphate (KMeth) or potassium gluconate (KGluc). Here we show immediate and gradual effects of these intracellular solutions on measurement of the AHP and basic membrane properties, and on the induction of AHP plasticity in CA1 pyramidal neurons from rat hippocampal slices. The AHP measured immediately after establishing whole-cell recordings was larger with KMeth than with KGluc. In general, the AHP in KMeth was comparable to the AHP measured in the perforated-patch configuration. However, KMeth induced time-dependent changes in the intrinsic membrane properties of CA1 pyramidal neurons. Specifically, input resistance progressively increased by 70% after 50 min; correspondingly, the current required to trigger an action potential and the fast afterdepolarization following action potentials gradually decreased by about 50%. Conversely, these measures were stable in KGluc. We also demonstrate that activity-dependent plasticity of the AHP occurs with physiologically relevant stimuli in KGluc. AHPs triggered with theta-burst firing every 30 s were progressively reduced, whereas AHPs elicited every 150 s were stable. Blockade of the apamin-sensitive AHP current (I(AHP)) was insufficient to block AHP plasticity, suggesting that plasticity is manifested through changes in the apamin-insensitive slow AHP current (sI(AHP)). These changes were observed in the presence of synaptic blockers, and therefore reflect changes in the intrinsic properties of the neurons. However, no AHP plasticity was observed using KMeth. In summary, these data show that KMeth produces time-dependent changes in basic membrane properties and prevents or obscures activity-dependent reduction of the AHP. In whole-cell recordings using KGluc, repetitive theta-burst firing induced AHP plasticity that mimics learning-related reduction in the AHP.