Article

Kv4 (A‐type) potassium currents in the mouse medial nucleus of the trapezoid body

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Abstract

Principal neurones of the mouse medial nucleus of the trapezoid body (MNTB) possess multiple voltage-gated potassium currents, including a transient outward current (or A-current), which is characterized here. The A-current exhibited rapid voltage-dependent inactivation and was half inactivated at resting membrane potentials. Following a hyperpolarizing pre-pulse to remove inactivation, the peak transient current was 1.07 nA at -17 mV. The pharmacological characteristics of this A-current were consistent with Kv4 subunits in expression studies; the A-current was resistant to block by tetraethylammonium and dendrotoxin-I but sensitive to millimolar concentrations of 4-aminopyridine and 5 microM hanatoxin. Immunohistochemistry confirmed that Kv4.3 sub-units are present in the MNTB. In a single-compartment model of an MNTB neurone, the A-current served to accelerate the decay of the initial action potentials in a stimulus train and suggested that removal of A-current steady-state inactivation could raise firing threshold for non-calyceal synaptic inputs. This A-type current was not observed in the rat.

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... The voltage dependence and the transient nature of the TEA isolated outward currents in VE and LOC neurons is highly compatible with the low voltage-activated A-type currents mediated by the Kv4 family of ion channels [4]. This ion current is mediated via the Kv4 channel family, which includes three poreforming a-subunits: Kv4.1, Kv4.2 and Kv4.3; of which Kv4.2 and Kv4.3 are specific for the brainstem [21], [33], [61]. In order to investigate if Kv4 subunits are expressed in VE neurons and, in that case which subtype, immunolabeling against ChAT, identifying the VE and LOC neurons, was combined with Kv4.2 and Kv4.3 antibodies in wild type animals. ...
... However, this protocol failed to produce any specific Kv4.3 staining in the mouse. In accordance with another study of Kv4immunolabeling in mice [33], the tissue preparation was switched from trans-cardial perfusion to a protocol using shock-freezing of the brain tissue. Presumably, this method better preserved antigenicity or opened up the cell membrane for better antibody-targeting of intracellular epitopes. ...
... The A current, carried by the Kv4 channels [4], is a rapidly activating and inactivating K + current [32]. I A is characteristically active at the resting membrane potential, usually demonstrated as a 'window-current' in the 270 to 250 mV range [33], [63], [70]. The large transient outward currents, with instantaneous activation and rapid inactivation upon depolarization, being active at resting membrane voltages in VE neurons, are highly compatible with I A . ...
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The functional role of efferent innervation of the vestibular end-organs in the inner ear remains elusive. This study provides the first physiological characterization of the cholinergic vestibular efferent (VE) neurons in the brainstem by utilizing a transgenic mouse model, expressing eGFP under a choline-acetyltransferase (ChAT)-locus spanning promoter in combination with targeted patch clamp recordings. The intrinsic electrical properties of the eGFP-positive VE neurons were compared to the properties of the lateral olivocochlear (LOC) brainstem neurons, which gives rise to efferent innervation of the cochlea. Both VE and the LOC neurons were marked by their negative resting membrane potential
... Kv3 channels speed AP repolarization, promoting short APs and high frequency firing (Wang et al. 1998;Rudy et al. 1999;Dodson & Forsythe, 2004). A-type potassium currents (I A ) mediated by Kv4 channels are also present in mouse MNTB neurons (Johnston et al. 2008). ...
... The recordings were made from animals aged P10-P14. The TEA and DTx-I-insensitive current has two distinct components, as shown in Fig. 1B: a fast activating transient A-type current (arrow) which has been characterized previously (Johnston et al. 2008) and a slowly activating delayed rectifier. ...
... Although many of the Kv7 channels (KCNQ/M-current) mediate slowly activating currents, they are all sensitive to low concentrations of linopirdine (Robbins, 2001;Lawrence et al. 2006;Vervaeke et al. 2006;Hu et al. 2007) and the slow outward current was unaffected by 50 μM linopirdine (n = 2, data not shown). The Kv4 family underlies A-type currents (Birnbaum et al. 2004) and Kv4.3 is responsible for the A-current observed here (Johnston et al. 2008) but A-currents were removed by voltage-dependent inactivation (see Fig. 2A). EAG/ERG/ELK (Kv10, 11 and 12) subfamilies were excluded from the present study as addition of E4031 (1 μM) to the internal patch solution and left the current unaffected (Gessner & Heinemann, 2003;Royer et al. 2005;Lamarca et al. 2006;Furlan et al. 2007). ...
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The medial nucleus of the trapezoid body (MNTB) is specialized for high frequency firing by expression of Kv3 channels, which minimize action potential (AP) duration, and Kv1 channels, which suppress multiple AP firing, during each calyceal giant EPSC. However, the outward K(+) current in MNTB neurons is dominated by another unidentified delayed rectifier. It has slow kinetics and a peak conductance of approximately 37 nS; it is half-activated at -9.2 +/- 2.1 mV and half-inactivated at -35.9 +/- 1.5 mV. It is blocked by several non-specific potassium channel antagonists including quinine (100 microm) and high concentrations of extracellular tetraethylammonium (TEA; IC(50) = 11.8 mM), but no specific antagonists were found. These characteristics are similar to recombinant Kv2-mediated currents. Quantitative RT-PCR showed that Kv2.2 mRNA was much more prevalent than Kv2.1 in the MNTB. A Kv2.2 antibody showed specific staining and Western blots confirmed that it recognized a protein approximately 110 kDa which was absent in brainstem tissue from a Kv2.2 knockout mouse. Confocal imaging showed that Kv2.2 was highly expressed in axon initial segments of MNTB neurons. In the absence of a specific antagonist, Hodgkin-Huxley modelling of voltage-gated conductances showed that Kv2.2 has a minor role during single APs (due to its slow activation) but assists recovery of voltage-gated sodium channels (Nav) from inactivation by hyperpolarizing interspike potentials during repetitive AP firing. Current-clamp recordings during high frequency firing and characterization of Nav inactivation confirmed this hypothesis. We conclude that Kv2.2-containing channels have a distinctive initial segment location and crucial function in maintaining AP amplitude by regulating the interspike potential during high frequency firing.
... Kv3 channels speed AP repolarization, promoting short APs and high frequency firing (Wang et al. 1998;Rudy et al. 1999;Dodson & Forsythe, 2004). A-type potassium currents (I A ) mediated by Kv4 channels are also present in mouse MNTB neurons (Johnston et al. 2008). ...
... The recordings were made from animals aged P10-P14. The TEA and DTx-I-insensitive current has two distinct components, as shown in Fig. 1B: a fast activating transient A-type current (arrow) which has been characterized previously (Johnston et al. 2008) and a slowly activating delayed rectifier. ...
... Although many of the Kv7 channels (KCNQ/M-current) mediate slowly activating currents, they are all sensitive to low concentrations of linopirdine (Robbins, 2001;Lawrence et al. 2006;Vervaeke et al. 2006;Hu et al. 2007) and the slow outward current was unaffected by 50 μM linopirdine (n = 2, data not shown). The Kv4 family underlies A-type currents (Birnbaum et al. 2004) and Kv4.3 is responsible for the A-current observed here (Johnston et al. 2008) but A-currents were removed by voltage-dependent inactivation (see Fig. 2A). EAG/ERG/ELK (Kv10, 11 and 12) subfamilies were excluded from the present study as addition of E4031 (1 μM) to the internal patch solution and left the current unaffected (Gessner & Heinemann, 2003;Royer et al. 2005;Lamarca et al. 2006;Furlan et al. 2007). ...
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This is the author's final draft of the version publsihed as Journal of Physiology, 2008, and can aslo be accessed via http://jp.physoc.org/cgi/content/abstract/jphysiol.2008.153734v1
... Previous in situ hybridization (ISH) experiments in rodent brain have shown that Kv4.1 staining only is present in the olfactory bulb and CA1 of the hippocampus (Serodio and Rudy, 1998;Fitzakerley et al., 2000). This has been confirmed with immunohistochemistry (IHC) in neurons of the medial nucleus of the trapezoid body (MNTB), which are functionally related to the neurons of interest and show a lack of Kv4.1 labelling (Johnston et al., 2008). This leaves Kv4.2 and Kv4.3 as possible candidates for causing the current in LOC and MOC neurons. ...
... Moreover, we also wanted to investigate if also the vestibular efferents express Kv4 channels and, in that case, which specific subunits that are present in these cells. The third member of the Kv4 subfamily, Kv4.1, has previously been shown not to be expressed in the areas of interest (Serodio and Rudy, 1998;Fitzakerley et al., 2000;Johnston et al., 2008) and was, therefore, not investigated. ...
... Principal cells of the SOC receive an ipsilateral excitatory input from the cochlear nucleus (CN) and a contralaterally driven feed-forward inhibitory input from the MNTB, which are used for extracting interaural intensity differences; an important cue for sound localization (Magnusson et al., 2008). Interestingly, we also found a strong expression of Kv4.3 in the rat MNTB (Fig.6C), which is in contradiction to the lack of transient outward currents in the rat MNTB observed by Johnston et al. (2008). ...
... As in the other rodent species, this specialization might be due to be expression of low voltage-activated potassium currents (mediated by Kv1 channels) which promote short latency transmission and minimize jitter as shown for the mouse (Brew et al., 2003;Gittelman and Tempel, 2006) and rat (Brew and Forsythe, 1995;Dodson et al., 2002). The mouse MNTB also expresses a rapidly inactivating low-voltage-activated current (A-current, mediated by Kv4 channels; (Brew et al., 2003;Johnston et al., 2008)), and the larger jitter observed in rat MNTB neurons is consistent with the lack of A-current in this species (Johnston et al., 2008). ...
... As in the other rodent species, this specialization might be due to be expression of low voltage-activated potassium currents (mediated by Kv1 channels) which promote short latency transmission and minimize jitter as shown for the mouse (Brew et al., 2003;Gittelman and Tempel, 2006) and rat (Brew and Forsythe, 1995;Dodson et al., 2002). The mouse MNTB also expresses a rapidly inactivating low-voltage-activated current (A-current, mediated by Kv4 channels; (Brew et al., 2003;Johnston et al., 2008)), and the larger jitter observed in rat MNTB neurons is consistent with the lack of A-current in this species (Johnston et al., 2008). ...
Article
Principal cells of the medial nucleus of the trapezoid body (MNTB) receive their excitatory input through large somatic terminals, the calyces of Held, which arise from axons of globular bushy cells located in the contralateral ventral cochlear nucleus. Discharges of MNTB neurons are characterized by high stimulus evoked firing rates, temporally precise onset responses, and a high degree of phase-locking to either pure tones or stimulus envelopes. Since the calyx of Held synapse is accessible to in vitro and to in vivo recordings, it serves as one of the most elaborate models for studying synaptic transmission in the mammalian brain. Although in such studies, the major emphasis is on synaptic physiology, the interpretation of the data will benefit from an understanding of the MNTB's contribution to auditory signal processing, including possible functional differences in different species. This implies the consideration of possible functional differences in different species. Here, we compare single unit recordings from MNTB principal cells in vivo in three different rodent species: gerbil, mouse and rat. Because of their good low-frequency hearing gerbils are often used in in vivo preparations, while mice and rats are predominantly used in slice preparations. We show that MNTB units in all three species exhibit high firing rates and precise onset-timing. Still there are species-specific specializations that might suggest the preferential use of one species over the others, depending on the scope of the respective investigation.
... Brain slices were prepared as described previously (Postlethwaite et al., 2007; Johnston et al., 2008a). Briefly, CBA/Ca mice or Lister Hooded rats aged P10–P19 were decapitated in accordance with the UK animals (Scientific Procedures) Act 1986 and the brain was removed in a slush of iced artificial CSF (aCSF) of composition (in mM) 250 sucrose, 2.5 KCl, 10 glucose, 1.25 NaH 2 PO 4 , 0.5 ascorbic acid, 26 NaHCO 3 , 4 MgCl 2 , 0.1 CaCl 2 , gassed with 95% O 2 /5% CO 2 (pH 7.4). ...
Article
Most current clamp studies trigger action potentials (APs) by step current injection through the recording electrode and assume that the resulting APs are essentially identical to those triggered by orthodromic synaptic inputs. However this assumption is not always valid, particularly when the synaptic conductance is of large magnitude and of close proximity to the axon initial segment. We addressed this question of similarity using the Calyx of Held/MNTB synapse; we compared APs evoked by long duration step current injections, short step current injections and orthodromic synaptic stimuli. Neither injected current protocol evoked APs that matched the evoked orthodromic AP waveform, showing differences in AP height, half-width and after-hyperpolarization. We postulated that this ‘error’ could arise from changes in the instantaneous conductance during the combined synaptic and AP waveforms, since the driving forces for the respective ionic currents are integrating and continually evolving over this time-course. We demonstrate that a simple Ohm's law manipulation of the EPSC waveform, which accounts for the evolving driving force on the synaptic conductance during the AP, produces waveforms that closely mimic those generated by physiological synaptic stimulation. This stimulation paradigm allows supra-threshold physiological stimulation (single stimuli or trains) without the variability caused by quantal fluctuation in transmitter release, and can be implemented without a specialised dynamic clamp system. Combined with pharmacological tools this method provides a reliable means to assess the physiological roles of postsynaptic ion channels without confounding affects from the presynaptic input.
... Channels containing Kv3 have high threshold and fast kinetics, enhancing rapid J Physiol 587.11 repolarisation, giving short duration APs and promoting high-frequency firing (Brew & Forsythe, 1995;Wang et al. 1998). There is also a small contribution from Kv4 (Johnston et al. 2008a), which acts to accelerate the initial AP time course during repetitive firing. Although these four families contribute the majority of outward potassium currents, other families may also make important contributions. ...
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The ionic basis of excitability requires identification and characterisation of expressed channels and their specific roles in native neurons. We have exploited principal neurons of the medial nucleus of the trapezoid body (MNTB) as a model system for examining voltage-gated K(+) channels, because of their known function and simple morphology. Here we show that channels of the ether-à-go-go-related gene family (ERG, Kv11; encoded by kcnh) complement Kv1 channels in regulating neuronal excitability around threshold voltages. Using whole-cell patch clamp from brainstem slices, the selective ERG antagonist E-4031 reduced action potential (AP) threshold and increased firing on depolarisation. In P12 mice, under voltage-clamp with elevated [K(+)](o) (20 mm), a slowly deactivating current was blocked by E-4031 or terfenadine (V(0.5,act) = -58.4 +/- 0.9 mV, V(0.5,inact) = -76.1 +/- 3.6 mV). Deactivation followed a double exponential time course (tau(slow) = 113.8 +/- 6.9 ms, tau(fast) = 33.2 +/- 3.8 ms at -110 mV, tau(fast) 46% peak amplitude). In P25 mice, deactivation was best fitted by a single exponential (tau(fast) = 46.8 +/- 5.8 ms at -110 mV). Quantitative RT-PCR showed that ERG1 and ERG3 were the predominant mRNAs and immunohistochemistry showed expression as somatic plasma membrane puncta on principal neurons. We conclude that ERG currents complement Kv1 currents in limiting AP firing at around threshold; ERG may have a particular role during periods of high activity when [K(+)](o) is elevated. These ERG currents suggest a potential link between auditory hyperexcitability and acoustic startle triggering of cardiac events in familial LQT2.
... The difference in P K 's provides an explanation for these results. The primary thrust of this review concerns the use of GHK normalization to obtain ion channel activation curves for the delayed rectifi er I K and for I A. A number of groups ( DeFazio and Moenter, 2002;Boland et al., 2003;Van Hoorick et al., 2003;Persson et al., 2005;Nakamura and Takahashi, 2007;Johnston et al., 2008;Dementieva et al., 2009;Sculptoreanu et al., 2009) have recently used the procedure outlined here and in a previous report from this laboratory (Clay, 2000) to obtain these results. One of the purposes of this review is to encourage others to do the same. ...
Article
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Potassium ion current in nerve membrane, I(K), has traditionally been described by I(K) = g(K)(V - E(K)), where g(K) is the K ion conductance, V is membrane potential and E(K) is the K(+) Nernst potential. This description has been unchallenged by most investigators in neuroscience since its introduction almost 60 years ago. The problem with the I(K) approximately (V - E(K)) proportionality is that it is inconsistent with the unequal distribution of K ions in the intra- and extracellular bathing media. Under physiological conditions the intracellular K(+) concentration is significantly higher than the extracellular concentration. Consequently, the slope conductance at potentials positive to E(K) cannot be the same as that for potentials negative to E(K), as the linear proportionality between I(K) and (V - E(K)) requires. Instead I(K) has a non-linear dependence on (V - E(K)) which is well described by the Goldman-Hodgkin-Katz equation. The implications of this result for K(+) channel gating and membrane excitability are reviewed in this report.
... First, the I A induced in NAergic A7 neurons displayed a voltage-independent inactivation rate, which distinguishes the I K mediated by Kv4 channels from those mediated by other K channels (Jerng et al., 2004a). Although small differences were noted, the half-points and the slopes of the activation/ inactivation curves, the time constants of inactivation, and the recovery from inactivation of the I A in NAergic neurons were all compatible with the values reported for Kv4.2 or Kv4.3 channels in other neurons in the CNS, including hippocampal pyramidal cells and interneruons (Martina et al., 1998), midbrain dopaminergic neurons (Liss et al., 2001), neurons in the medial nucleus of the trapezoid body (Johnston et al., 2008), and cerebellar granule cells (Shibata et al., 2000). In these studies, the fact that the I A were mediated by Kv4.3 or Kv4.2 channels was proved by experiments that either showed a positive correlation between the current density of the I A and the amount of Kv4 channel transcription or block of the I A by specific knock down of Kv4 channel function. ...
Article
We investigated voltage-dependent K(+) currents (I(K)) in noradrenergic (NAergic) A7 neurons. The I(K) evoked consisted of A-type I(K) (I(A)), which had the characteristics of a low threshold for activation (approximately -50 mV), fast activation/inactivation, and rapid recovery from inactivation. Since the I(A) were blocked by heteropodatoxin-2 (Hptx-2), a specific Kv4 channel blocker, and the NAergic A7 neurons were shown to be reactive with antibodies against Kv4.1/Kv4.3 channel proteins, we conclude that the I(A) evoked in NAergic neurons are mediated by Kv4.1/Kv4.3 channels. I(A) were also evoked using voltage commands of a single action potential (AP), a subthreshold voltage change between two consecutive APs, or excitatory postsynaptic potential (EPSP) activity recorded in current-clamp mode (CCM). Blockade of the I(A) by 4-AP, a broad spectrum I(A) blocker, or by Hptx-2 increased the half-width and spontaneous firing of APs and reduced the amount of synaptic drive needed to elicit APs in CCM, showing that the I(A) play important roles in regulating the shape and firing frequency of APs and in synaptic integration in NAergic A7 neurons. Since these neurons are the principal projection neurons to the dorsal horn of the spinal cord, these results also suggest roles for Kv4.1/4.3 channels in descending NAergic pain regulation.
... A similar role has been proposed for sodium-activated K + (IK Na ) channels (Yang et al. 2007) although IK Na may not contribute under all experimental conditions (Johnston et al. 2008a). Other potassium conductances mediated by HCN, ERG and Kv4 channels (Johnston et al. 2008b) are present, but with relatively small conductances and further work is required to understand their various specific and complementary roles in MNTB function. Potassium currents are crucial for MNTB neurons to maintain high firing rates, avoiding aberrant firing, and to permit sustained longer periods of firing. ...
Article
Full-text available
In this review we take a physiological perspective on the role of voltage-gated potassium channels in an identified neuron in the auditory brainstem. The large number of KCN genes for potassium channel subunits and the heterogeneity of the subunit combination into K(+) channels make identification of native conductances especially difficult. We provide a general pharmacological and biophysical profile to help identify the common voltage-gated K(+) channel families in a neuron. Then we consider the physiological role of each of these conductances from the perspective of the principal neuron in the medial nucleus of the trapezoid body (MNTB). The MNTB is an inverting relay, converting excitation generated by sound from one cochlea into inhibition of brainstem nuclei on the opposite side of the brain; this information is crucial for binaural comparisons and sound localization. The important features of MNTB action potential (AP) firing are inferred from its inhibitory projections to four key target nuclei involved in sound localization (which is the foundation of auditory scene analysis in higher brain centres). These are: the medial superior olive (MSO), the lateral superior olive (LSO), the superior paraolivary nucleus (SPN) and the nuclei of the lateral lemniscus (NLL). The Kv families represented in the MNTB each have a distinct role: Kv1 raises AP firing threshold; Kv2 influences AP repolarization and hyperpolarizes the inter-AP membrane potential during high frequency firing; and Kv3 accelerates AP repolarization. These actions are considered in terms of fidelity of transmission, AP duration, firing rates and temporal jitter. An emerging theme is activity-dependent phosphorylation of Kv channel activity and suggests that intracellular signalling has a dynamic role in refining neuronal excitability and homeostasis.
... That portion of the model is not a factor in type 3 behaviour (simulations not shown). Several groups have recently used the GHK normalization procedure described in this study and in an earlier report (Clay 2000) to obtain K C channel activation curves (DeFazio & Moenter 2002; Boland et al. 2003; Van Hoorick et al. 2003; Persson et al. 2005; Nakamura & Takahashi 2007; Johnston et al. 2008). A far larger number of reports have used normalization with (VKE K ). ...
Article
The Hodgkin and Huxley (HH) model predicts sustained repetitive firing of nerve action potentials for a suprathreshold depolarizing current pulse for as long as the pulse is applied (type 2 excitability). Squid giant axons, the preparation for which the model was intended, fire only once at the beginning of the pulse (type 3 behaviour). This discrepancy between the theory and experiments can be removed by modifying a single parameter in the HH equations for the K+ current as determined from the analysis in this paper. K+ currents in general have been described by IK=gK(V-EK), where gK is the membrane's K+ current conductance and EK is the K+ Nernst potential. However, IK has a nonlinear dependence on (V-EK) well described by the Goldman-Hodgkin-Katz equation that determines the voltage dependence of gK. This experimental finding is the basis for the modification in the HH equations describing type 3 behaviour. Our analysis may have broad significance given the use of IK=gK(V-EK) to describe K+ currents in a wide variety of biological preparations.
... Several other types of potassium channels are expressed in the MNTB of different species and at different times in development. These include Kv4.3 subunits, which have been reported in the MNTB of mice but not that of rats (Johnston et al., 2008a). These subunits generate rapidly inactivating "A-type" potassium currents, which could potentially influence the timing of neuronal responses, but their precise role is not understood. ...
Article
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The extraction and localization of an auditory stimulus of interest from among multiple other sounds, as in the ‘cocktail-party’ situation, requires neurons in auditory brainstem nuclei to encode the timing, frequency, and intensity of sounds with high fidelity, and to compare inputs coming from the two cochleae. Accurate localization of sounds requires certain neurons to fire at high rates with high temporal accuracy, a process that depends heavily on their intrinsic electrical properties. Studies have shown that the membrane properties of auditory brainstem neurons, particularly their potassium currents, are not fixed but are modulated in response to changes in the auditory environment. Here, we review work focusing on how such modulation of potassium channels is critical to shaping the firing pattern and accuracy of these neurons. We describe how insights into the role of specific channels have come from human gene mutations that impair localization of sounds in space. We also review how short-term and long-term modulation of these channels maximizes the extraction of auditory information, and how errors in the regulation of these channels contribute to deficits in decoding complex auditory information.
Chapter
Hodgkin-Huxley model (HH model) qualitatively describes the generation of the action potential of squid giant axons. The resting state or oscillatory phase state of the membrane potential (voltage) in the HH model depends mainly on applied stimulus (external currents) to neurons. The firing of the action potential in neurons depends upon the depolarization or repolarization of ions. The probability of channel gates (to be open or close) determines the movement of ions across the cell membrane. The term of K+ ionic currents (related to several activation gates) in external current contains exponential power 4 in HH model in which we propose a modified HH model by considering the higher power (5 and 6) of K activation in potassium ionic currents and studied the behavior of all three models comparatively. The modified HH model with a higher power of potassium activation reached resting-state sooner and gains stability (after oscillatory) at a high external current. The qualitative behavior of the modified model (with the higher exponential power) is different as there is a shifting of Hopf bifurcation points in comparison with the original HH model. Moreover, a larger periodic region was observed in most of the parameter phase spaces (external current I versus parameters) except against the Na conductance and Na potential. The modified HH model which determines that higher power of K activation is more significant for action potential in neurons.
Article
To perform auditory tasks such as sound localization in the space, auditory neurons in the brain must distinguish sub-millisecond temporal differences in signals from two ears. Such high temporal resolution is possible when each neuron in the ascending auditory pathway fires brief action potential at very accurate timing. Various pre- and postsynaptic machineries ensuring such high temporal precision of auditory synaptic transmission have been identified. Of particular, in this review, the role of K(+) channels in shortening the duration of synaptic potentials will be discussed. First, the contribution of K(+) channels to AP firing of general auditory neurons will be discussed. Then, the focus will be moved to the inner hair cell (IHC)-auditory afferent nerve fiber (ANF) synapses, the first synapses of ascending auditory pathway. Molecular and immunohistological techniques have revealed various K(+) channels in the cell bodies and their processes of ANFs. Since the development of patch-clamp recordings from the ANF dendrites in 2002, it became possible to monitor the IHC-ANF synaptic transmission in greater detail. As revealed in brain auditory synapses, several different K(+) channels appear to participate in reducing the duration of synaptic potentials at the IHC-ANF synapses. In addition, K(+) channels at the ANF dendrites might act as potential targets of efferent feedback from the brain. The hypothesis is that, upon loud sound exposure, efferent neurotransmitters released onto the ANF dendrites activate certain K(+) channels and prevent excitotoxicity of ANFs. Therefore, K(+) channels of the ANF dendrites might provide potential sites of pharmacological actions to prevent noise-induced hearing loss.
Article
This investigation compared the development of neuronal excitability in the ventral nucleus of the trapezoid body (VNTB) between two strains of mice with differing progression rates for age-related hearing loss. In contrast to CBA/Ca (CBA) mice, the C57BL/6J (C57) strain are subject to hearing loss from a younger age and are more prone to damage from sound over-exposure. Higher firing rates in the medial olivocochlear system (MOC) are associated with protection from loud sounds and these cells are located in the VNTB. We postulated that reduced neuronal firing of the MOC in C57 mice could contribute to hearing loss in this strain by reducing efferent protection. Whole cell patch clamp was used to compare the electrical properties of VNTB neurons from the two strains initially in two age groups: before and after hearing onset at ∼ P9 and ∼P16, respectively. Prior to hearing onset VNTB neurons electrophysiological properties were identical in both strains, but started to diverge after hearing onset. One week after hearing onset VNTB neurons of C57 mice had larger amplitude action potentials but in contrast to CBA mice, their waveform failed to accelerate with increasing age, consistent with the faster inactivation of voltage-gated potassium currents in C57 VNTB neurons. The lower frequency action potential firing of C57 VNTB neurons at P16 was maintained to P28, indicating that this change was not a developmental delay. We conclude that C57 VNTB neurons fire at lower frequencies than in the CBA strain, supporting the hypothesis that reduced MOC firing could contribute to the greater hearing loss of the C57 strain.
Article
The auditory part of the brainstem is composed of several nuclei specialized in the computation of the different spectral and temporal features of the sound before it reaches the higher auditory regions. There are a high diversity of neuronal types in these nuclei, many with remarkable electrophysiological and synaptic properties unique to these structures. This diversity reflects specializations necessary to process the different auditory signals in order to extract precisely the acoustic information necessary for the auditory perception by the animal. Low threshold Kv1 channels and HCN channels are expressed in neurons that use timing clues for auditory processing, like bushy and octopus cells, in order to restrict action potential firing and reduce input resistance and membrane time constant. Kv3 channels allow principal neurons of the MNTB and pyramidal DCN neurons to fire fast trains of action potentials. Calcium channels on cartwheel DCN neurons produce complex spikes characteristic of these neurons. Calyceal synapses compensate the low input resistance of bushy and principal neurons of the MNTB by releasing hundreds of glutamate vesicles resulting in large EPSCs acting in fast ionotropic glutamate receptors, in order to reduce temporal summation of synaptic potentials, allowing more precise correspondence of pre- and post-synaptic potentials, and phase-locking. Pre-synaptic calyceal sodium channels have fast recovery from inactivation allowing extremely fast trains of action potential firing, and persistent sodium channels produce spontaneous activity of fusiform neurons at rest, which expands the dynamic range of these neurons. The unique combinations of different ion channels, ionotropic receptors and synaptic structures create a unique functional diversity of neurons extremely adapted to their complex functions in the auditory processing.
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The largest biomass of channel proteins is located in unicellular organisms and bacteria that have no organs. However, orchestrated bidirectional ionic currents across the cell membrane via the channels are important for the functioning of organs of organisms, and equally concern both fauna or flora. Several ion channels are activated in the course of action potentials. One of the hallmarks of voltage-dependent channels is a 'tail current' - deactivation as observed after prior and sufficient activation predominantly at more depolarized potentials e.g. for Kv while upon hyperpolarization for HCN α subunits. Tail current also reflects the timing of channel closure that is initiated upon termination of stimuli. Finally, deactivation of currents during repolarization could be a selective estimate for given channel as in case of HERG, if dedicated long and more depolarized 'tail pulse' is used. Since from a holding potential of e.g. -70 mV are often a family of outward K+ currents comprising IA and IK are simultaneously activated in native cells.
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Temporal coding precision of bushy cells in the ventral cochlear nucleus (VCN), critical for sound localization and communication, depends on the generation of rapid and temporally precise action potentials (APs). Voltage-gated potassium (Kv) channels are critically involved in this. The bushy cells in rat VCN express Kv1.1, 1.2, 1.3, 1.6, 3.1, 4.2 and 4.3 subunits. The Kv1.1 subunit contributes to the generation of a temporally precise single AP. However, the understanding of the functions of other Kv subunits expressed in the bushy cells is limited. Here, we investigated the functional diversity of Kv subunits concerning their contributions to temporal coding. We characterized the electrophysiological properties of the Kv channels with different subunits using whole-cell patch-clamp recording and pharmacological methods. The neuronal firing pattern changed from single to multiple APs only when the Kv1.1 subunit was blocked. The Kv subunits, including the Kv1.1, 1.2, 1.6 or 3.1, were involved in enhancing temporal coding by lowering membrane excitability, shortening AP latencies, reducing jitter and regulating AP kinetics. Meanwhile, all the Kv subunits contributed to rapid repolarization and sharpening peaks by narrowing half-width and accelerating fall rate, while the Kv1.1 subunit also affected the depolarization of AP. The Kv1.1, 1.2 and 1.6 subunits endowed bushy cells with a rapid time constant and a low input resistance of membrane for enhancing spike timing precision. The present results indicate that the Kv channels differentially affect intrinsic membrane properties to optimize the generation of rapid and reliable APs for temporal coding.
Chapter
Transient outward potassium currents were first described nearly 60 years ago, since then major strides have been made in understanding their molecular basis and physiological roles. From the large family of voltage-gated potassium channels members of 3 subfamilies can produce such fast-inactivating A-type potassium currents. Each subfamily gives rise to currents with distinct biophysical properties and pharmacological profiles and a simple workflow is provided to aid the identification of channels mediating A-type currents in native cells. Their unique properties and regulation enable A-type K+ channels to perform varied roles in excitable cells including repolarisation of the cardiac action potential, controlling spike and synaptic timing, regulating dendritic integration and long-term potentiation as well as being a locus of neural plasticity.
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We develop a new class of distributions, namely, the exponentiated half logistic-Topp-Leone-G power series (EHL-TL-GPS) class of distributions. We present some special classes in the proposed distribution. Structural properties were also derived including moments, entropy and maximum likelihood estimates. We conducted a simulation study to evaluate the consistency of the maximum likelihood estimates. We also present two real data examples to illustrate the applicability of the new class of distributions. The proposed model performs better than several non-nested models on selected data sets.
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1. We have used whole-cell and single-channel recording to study the transient outward potassium current (A-current) of rat locus coeruleus neurones grown in tissue culture. The A-current was largely inactivated at the resting potential, but could be activated from sufficiently negative holding potentials during steps positive to -50 mV. The current was sensitive to 4-aminopyridine. Another slowly activating, sustained current was similar to a delayed rectifier. 2. In the on-cell configuration the unitary conductance of channels carrying A-current was 40.9 +/- 2.2 pS (n = 6) with high external potassium (140 mM) and 14.8 +/- 1.4 pS (n = 11) with 3 mM [K+]o. The unitary current-voltage relation was not linear, but had a negative slope at very positive voltages in 3 mM [K+]o. The reversal potential changed with [K]o as expected for a K+ channel. 3. The open state probability of A-current channels was voltage dependent, reaching a peak of 0.78 +/- 0.17 (seven patches). The relationships between both activation and inactivation and membrane potential were well fitted by Boltzmann expressions. Activation was half-maximum at a potential 71.9 +/- 11.8 mV (n = 4) positive to the resting potential (approximately -61 mV). Inactivation was half-complete 29.4 +/- 3.8 mV (n = 4) negative to the resting potential. There was evidence from runs analysis for slow inactivation of channels. 4. Channels showed frequent visits to substates, the most readily identifiable of which had an amplitude 0.55 +/- 0.04 (n = 5) of the fully open state. Other substates had amplitudes of around 0.25 and 0.75. Occupancy of substates was greater at negative membrane potentials. 5. A preliminary analysis of kinetic behaviour, treating visits to substates as openings, shows that open times are distributed as a single exponential. The open time was 16.2 ms (n = 4) at a voltage 100 mV positive to the resting potential, increasing with further depolarization. Closed times are distributed as the sum of three or four exponentials. First latency distributions are strongly voltage dependent and show a delay, giving a sigmoidal rise to the distribution. Increasing temperature increased unitary current and reduced mean open time. 6. The mechanism of the rectification seen in the unitary current-voltage relationship was examined using excised, inside-out patches.(ABSTRACT TRUNCATED AT 400 WORDS)
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1. Membrane current-voltage relationships were investigated under voltage clamp conditions in isolated neural somata of marine gastropods.2. Step depolarizations from the resting potential produce an initial inward current followed by a delayed outward current.3. Inward current appears to be carried by both sodium and calcium ions and displays time and voltage dependent properties similar to those of other excitable membranes.4. Activation and deactivation of the delayed outward current follow a more complicated time course than that of a single exponential raised to a power but can be fitted by the product of two exponential functions.5. Delayed outward current inactivation proceeds with a time constant which decreases as membrane voltage is made more positive. The steady-state levels of inactivation as a function of membrane voltage are related by an S-shaped curve similar to that for K inactivation in squid giant axon.
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The Kv2.1 voltage-activated K+ channel, a Shab-related K+ channel isolated from rat brain, is insensitive to previously identified peptide inhibitors. We have isolated two peptides from the venom of a Chilean tarantula, G. spatulata, that inhibit the Kv2.1 K+ channel. The two peptides, hanatoxin1 (HaTx1) and hanatoxin2 (HaTx2) are unrelated in primary sequence to other K+ channel inhibitors. The activity of HaTx was verified by synthesizing it in a bacterial expression system. The concentration dependence for both the degree of inhibition at equilibrium (Kd = 42 nM) and the kinetics of inhibition (kon = 3.7 x 10(4) M-1s-1; koff = 1.3 x 10(-3) s-1), are consistent with a bimolecular reaction between HaTx and the Kv2.1 K+ channel. Shaker-related, Shaw-related, and eag K+ channels were relatively insensitive to HaTx, whereas a Shal-related K+ channel was sensitive. Regions outside the scorpion toxin binding site (S5-S6 linker) determine sensitivity to HaTx. HaTx introduces a new class of K+ channel inhibitors that will be useful probes for studying K+ channel structure and function.
Article
1. Whole-cell voltage-clamp techniques were used to study voltage-activated transient potassium currents in a large sample (n = 143) of granule cells (GrC) from rat cerebellar slices. Tetrodotoxin (TTX; 0.1 microM) was used to block sodium currents, while calcium current was too small to be seen under ordinary conditions. 2. Depolarizing pulses from -50 mV evoked a slow, sustained outward current, developing with a time constant of 10 ms, inactivating over a time scale of seconds and which could be suppressed by 20 mM tetraethylammonium (TEA). By preventing the Ca2+ inflow, this slow outward current could be further separated into a Ca(2+)-dependent and a Ca(2+)-independent component. 3. After conditioning hyperpolarizations to potentials negative to -60 mV, depolarizations elicited transient outward current, peaking in 1-2 ms and inactivating rapidly (approximately 10 ms at 20 degrees C), showing the overall kinetic characteristics of the A-current (IA). The current activated following third-order kinetics and showed a maximal conductance of 12 nS per cell, corresponding to a normalized conductance of 3.8 nS/pF. 4. IA was insensitive to TEA and to the Ca(2+)-channel blockers. 4-Aminopyridine (4-AP) reduced the A-current amplitude by approximately 20%, and the delayed outward currents by > 80%. 5. Voltage-dependent steady-state inactivation of peak IA was described by a Boltzmann function with a slope factor of 8.4 mV and half-inactivation occurring at -78.8 mV. Activation of IA was characterized by a Boltzmann curve with the midpoint at -46.7 mV and with a slope factor of 19.8 mV. 6. IA activation and inactivation was best fitted by the Hodgkin-Huxley m3h formalism. The rate of activation, tau a, was voltage-dependent, and had values ranging from 0.55 ms at -40 mV to 0.2 ms at +50 mV. Double-pulse experiment showed that development and removal of inactivation followed a single-exponential time course; the inactivation time constant, tau ha, was markedly voltage-dependent and ranged from approximately 10 ms at -40 and -100 mV and 70 ms at -70 mV. 7. A set of continuous equations has been developed describing the voltage-dependence of both the steady-state and time constant of activation and inactivation processes, allowing a satisfactory numerical reconstruction of the A-current over the physiologically significant membrane voltage range.(ABSTRACT TRUNCATED AT 400 WORDS)
Article
Mutagenesis experiments on voltage-gated K+ channels have suggested that the ion-selective pore is comprised mostly of H5 segments. To see whether regions outside of the H5 segment might also contribute to the pore structure, we have studied the effect of single amino acid substitutions in the segment that connects the S4 and S5 putative transmembrane segments (S4-S5 loop) on various permeation properties of Shaker K+ channels. Mutations in the S4-S5 loop alter the Rb+ selectivity, the single-channel K+ and Rb+ conductances, and the sensitivity to open channel block produced by intracellular tetraethylammonium ion, Ba2+, and Mg2+. The block of Shaker K+ channels by intracellular Mg2+ is surprising, but is reminiscent of the internal Mg2+ blockade of inward rectifier K+ channels. The results suggest that the S4-S5 loop constitutes part of the ion-selective pore. Thus, the S4-S5 loop and the H5 segment are likely to contribute to the long pore characteristic of voltage-gated K+ channels.
Article
1. The contribution of voltage-activated outward potassium currents to membrane excitability of neurons in the rat's dorsal nucleus of the lateral lemniscus (DNLL) was studied in a brain slice preparation using whole cell patch-clamp and intracellular recordings. Voltage-clamp methods and pharmacological manipulations were used to examine the currents regulating membrane dynamics in DNLL. 2. A delayed sustained outward current was evoked by applying depolarizing voltage steps across the cell membrane from a holding potential of -50 mV. An additional transient outward current was evoked when the depolarizing steps were preceded by a hyperpolarizing prepulse of -110 or -120 mV. 3. The transient outward current peaked within 6.8 ms of the onset of a depolarizing pulse. It decayed with a time constant of 12.3 ms for a 60-mV depolarizing voltage shift. Half-inactivation of this current occurred at -81.3 mV. The time constant for removal of the inactivation was 17.4 ms. The transient current had a high sensitivity to 4-aminopyridine (4-AP). 4. The sustained current was activated more slowly and was more sensitive to tetraethylammonium (TEA) than the transient current. The sustained current had both Ca2+-dependent and Ca2+-independent components. The Ca2+-dependent portion emerged at potentials of about -35 mV and was activated fully at +10 mV. The Ca2+-independent component was activated at potentials more positive than -40 mV and increased in magnitude with further depolarization. Inactivation of the Ca2+-independent component was voltage dependent. Also, TEA suppressed the Ca2+-independent compound. 5. The transient current in DNLL neurons closely resembled the A current (IA) described for hippocampal and other neurons in both kinetics and pharmacology. The Ca2+-independent component of the sustained current resembled the K current (IK) described for other neurons in both its properties of activation and inactivation and its pharmacology. 6. The outward current of some DNLL neurons was found to contain a dendrotoxin-sensitive component. This component reached its peak at 6.8 ms and had voltage-sensitive time constants of decay of 25.5 and 8.5 ms with voltage steps of 40 and 60 mV, respectively. 7. Application of 4-AP and TEA markedly prolonged the spike width, abolished the fast component of the after hyperpolarization and depolarized the cell membrane. Also, the number of action potentials produced by positive current injection increased under the influence of 4-AP and TEA. Membrane excitability and spike repolarization were dependent on both 4-AP-sensitive transient and TEA-sensitive sustained currents. 8. Neurons in DNLL typically exhibit a steady discharge of action potentials in response to sustained membrane depolarization. The rate and temporal pattern of production of action potentials in these cells are determined by the combination of transient and sustained potassium channels.
Article
The nonpeptide agent CP-339,818 (1-benzyl-4-pentylimino-1,4-dihydroquinoline) and two analogs (CP-393,223 and CP-394,322) that differ only with respect to the type of substituent at the N1 position, potently blocked the Kv1.3 channel in T lymphocytes. A fourth compound (CP-393,224), which has a smaller and less-lipophilic group at N1, was 100-200-fold less potent, suggesting that a large lipophilic group at this position is necessary for drug activity. CP-339,818 blocked Kv1.3 from the outside with a IC50 value of approximately 200 nM and 1:1 stoichiometry and competitively inhibited 125I-charybdotoxin from binding to the external vestibule of Kv1.3. This drug inhibited Kv1.3 in a use-dependent manner by preferentially blocking the C-type inactivated state of the channel. CP-339,818 was a significantly less potent blocker of Kv1.1, Kv1.2, Kv1.5, Kv1.6, Kv3.1-4, and Kv4.2; the only exception was Kv1.4, a cardiac and neuronal A-type K+ channel. CP-339,818 had no effect on two other T cell channels (I(CRAC) and intermediate-conductance K(Ca)) implicated in T cell mitogenesis. This drug suppresses human T cell activation, suggesting that blockade of Kv1.3 alone is sufficient to inhibit this process.
Article
Electrical properties of cochlear efferent (olivocochlear) neurons were investigated with the use of the whole cell patch recording technique in slice preparations of the neonatal rat (postnatal days 5-11). Lateral and medial olivocochlear (LOC and MOC, respectively) neurons were retrogradely labeled with a fluorescent tracer injected into the cochlea. Stained neurons were identified under a fluorescence microscope, and they were subjected to whole cell recording. LOC and MOC neurons showed different electrophysiological properties. Both showed spike trains of tonic pattern in response to injection of depolarizing current pulses at the resting membrane potential (-60 to -70 mV). However, when the membrane was slightly hyperpolarized (-72 to -76 mV), LOC neurons showed spike trains with a long first interspike interval (ISI), whereas MOC neurons showed spike trains with a long latency to the first spike. Extracellular application of 4-aminopyridine (4-AP; 0.5-2 mM) shortened these ISIs and latencies. In voltage-clamp experiments, two transient outward currents with different (fast and slow) decay kinetics were observed in LOC neurons. The fast outward current (I(A-LOC)) was inactivated by the preceding depolarization, and decayed with a time constant (tau) of 86 ms (at 0 mV). The preceding potential, which reduced the current size to the half-maximum (V1/2), was -72 mV. The slow current (I(KD)) decayed with a tau of 853 ms (at 0 mV). I(A-LOC) was sensitive to 4-AP (2 mM), and was less sensitive to tetraethylammonium chloride (TEA; 20 mM). I(KD) was partially blocked by TEA (20 mM), but was insensitive to 4-AP (2 mM). The recovery from inactivation of I(A-LOC) was time dependent with a time constant (tau(rec)) of 32 ms at -90 mV. MOC neurons also showed a transient outward current that consisted of a single transient component (I(A-MOC)) with a steady outward current. I(A-MOC) was inactivated by the preceding depolarization. Decay tau of I(A-MOC) was 33 ms (at 0 mV), and V1/2 was -75 mV. I(A-MOC) was sensitive to 4-AP (0.5-1 mM). The time-dependent recovery from inactivation of I(A-MOC) was faster than that of I(A-LOC), and tau(rec) was 15 ms at -90 mV. The different kinetics of transient outward currents between LOC and MOC neurons seems to be responsible for the difference in firing properties of these two neurons.
Article
The mammalian Kv4 gene subfamily and its Drosophila Shal counterpart encode proteins that form fast inactivating K+ channels that activate and inactivate at subthreshold potentials and recover from inactivation at a faster rate than other inactivating Kv channels. Taken together, the properties of Kv4 channels compare best with those of low-voltage activating "A-currents" present in the neuronal somatodendritic compartment and widely reported across several types of central and peripheral neurons, as well as the (Ca2+-independent) transient outward potassium conductance of heart cells (Ito). Three distinct genes have been identified that encode mammalian Shal homologs (Kv4. 1, Kv4.2, and Kv4.3), of which the latter two are abundant in rat adult brain and heart tissues. The distribution in the adult rat brain of the mRNA transcripts encoding the three known Kv4 subunits was studied by in situ hybridization histochemistry. Kv4.1 signals are very faint, suggesting that Kv4.1 mRNAs are expressed at very low levels, but Kv4.2 and Kv4.3 transcripts appear to be abundant and each produces a unique pattern of expression. Although there is overlap expression of Kv4.2 and Kv4.3 transcripts in several neuronal populations, the dominant feature is one of differential, and sometimes reciprocal expression. For example, Kv4.2 transcripts are the predominant form in the caudate-putamen, pontine nucleus and several nuclei in the medula, whereas the substantia nigra pars compacta, the restrosplenial cortex, the superior colliculus, the raphe, and the amygdala express mainly Kv4.3. Some brain structures contain both Kv4.2 and Kv4.3 mRNAs but each dominates in distinct neuronal subpopulations. For example, in the olfactory bulb Kv4.2 dominates in granule cells and Kv4.3 in periglomerular cells. In the hippocampus Kv4.2 is the most abundant isoform in CA1 pyramidal cells, whereas only Kv4.3 is expressed in interneurons. Both are abundant in CA2-CA3 pyramidal cells and in granule cells of the dentate gyrus, which also express Kv4.1. In the dorsal thalamus strong Kv4.3 signals are seen in several lateral nuclei, whereas medial nuclei express Kv4.2 and Kv4.3 at moderate to low levels. In the cerebellum Kv4.3, but not Kv4.2, is expressed in Purkinje cells and molecular layer interneurons. In the cerebellar granule cell layer, the reciprocity between Kv4.2 and Kv4.3 is observed in subregions of the same neuronal population. In fact, the distribution of Kv4 channel transcripts in the cerebellum defines a new pattern of compartmentation of the cerebellar cortex and the first one involving molecules directly involved in signal processing.
Article
1. Using the whole-cell recording mode we have characterized two non-conducting states in mammalian Shaker-related voltage-gated K+ channels induced by the removal of extracellular potassium, K+o. 2. In the absence of K+o, current through Kv1.4 was almost completely abolished due to the presence of a charged lysine residue at position 533 at the entrance to the pore. Removal of K+o had a similar effect on current through Kv1.3 when the histidine at the homologous position (H404) was protonated (pH 6.0). Channels containing uncharged residues at the corresponding position (Kv1.1: Y; Kv1.2: V) did not exhibit this behaviour. 3. To characterize the nature of the interaction between Kv1.3 and K+o concentration ([K+]o), we replaced H404 with amino acids of different character, size and charge. Substitution of hydrophobic residues (A, V and L) either in all four subunits or in only two subunits in the tetramer made the channel insensitive to the removal of K+o, possibly by stabilizing the channel complex. Replacement of H404 with the charged residue arginine, or the polar residue asparagine, enhanced the sensitivity of the channel to 0 mM K+o, possibly by making the channel unstable in the absence of K+o. Mutation at a neighbouring position (400) had a similar effect. 4. The effect of removing K+o on current amplitude does not seem to be correlated with the rate of C-type inactivation since the slowly inactivating G380F mutant channel exhibited a similar [K+]o dependence as the wild-type Kv1.3 channel. 5. CP-339,818, a drug that recognizes only the inactivated conformation of Kv1.3, could not block current in the absence of K+o unless the channels were inactivated through depolarizing pulses. 6. We conclude that removal of K+o induces the Kv1.3 channel to transition to a non-conducting 'closed' state which can switch into a non-conducting 'inactivated' state upon depolarization.
Article
1. Using a combination of patch-clamp, in situ hybridization and computer simulation techniques, we have analysed the contribution of potassium channels to the ability of a subset of mouse auditory neurones to fire at high frequencies. 2. Voltage-clamp recordings from the principal neurones of the medial nucleus of the trapezoid body (MNTB) revealed a low-threshold dendrotoxin (DTX)-sensitive current (ILT) and a high-threshold DTX-insensitive current (IHT). 3. IHT displayed rapid activation and deactivation kinetics, and was selectively blocked by a low concentration of tetraethylammonium (TEA; 1 mM). 4. The physiological and pharmacological properties of IHT very closely matched those of the Shaw family potassium channel Kv3.1 stably expressed in a CHO cell line. 5. An mRNA probe corresponding to the C-terminus of the Kv3.1 channel strongly labelled MNTB neurones, suggesting that this channel is expressed in these neurones. 6. TEA did not alter the ability of MNTB neurones to follow stimulation up to 200 Hz, but specifically reduced their ability to follow higher frequency impulses. 7. A computer simulation, using a model cell in which an outward current with the kinetics and voltage dependence of the Kv3.1 channel was incorporated, also confirmed that the Kv3.1- like current is essential for cells to respond to a sustained train of high-frequency stimuli. 8. We conclude that in mouse MNTB neurones the Kv3.1 channel contributes to the ability of these cells to lock their firing to high-frequency inputs.
Article
K+ channel principal subunits are by far the largest and most diverse of the ion channels. This diversity originates partly from the large number of genes coding for K+ channel principal subunits, but also from other processes such as alternative splicing, generating multiple mRNA transcripts from a single gene, heteromeric assembly of different principal subunits, as well as possible RNA editing and posttranslational modifications. In this chapter, we attempt to give an overview (mostly in tabular format) of the different genes coding for K+ channel principal and accessory subunits and their genealogical relationships. We discuss the possible correlation of different principal subunits with native K+ channels, the biophysical and pharmacological properties of channels formed when principal subunits are expressed in heterologous expression systems, and their patterns of tissue expression. In addition, we devote a section to describing how diversity of K+ channels can be conferred by heteromultimer formation, accessory subunits, alternative splicing, RNA editing and posttranslational modifications. We trust that this collection of facts will be of use to those attempting to compare the properties of new subunits to the properties of others already known or to those interested in a comparison between native channels and cloned candidates.
Article
Potassium ion channels are generally believed to have current-voltage (IV) relations which are linearly related to driving force ( V - E(K)), where V is membrane potential and E(K) is the potassium ion equilibrium potential. Consequently, activation curves for K+ channels have often been measured by normalizing voltage-clamp families of macroscopic K+ currents with (V - E(K)), where V is the potential of each successive step in the voltage clamp sequence. However, the IV relation for many types of K+ channels actually has a non-linear dependence upon driving force which is well described by the Goldman-Hodgkin-Katz relation. When the GHK dependence on (V - E(K)) is used in the normalization procedure, a very different voltage dependence of the activation curve is obtained which may more accurately reflect this feature of channel gating. Novel insights into the voltage dependence of the rapidly inactivating I(A) channels Kv1.4 and Kv4.2 have been obtained when this procedure was applied to recently published results.
Article
NEURON is a simulation environment for models of individual neurons and networks of neurons that are closely linked to experimental data. NEURON provides tools for conveniently constructing, exercising, and managing models, so that special expertise in numerical methods or programming is not required for its productive use. This article describes two tools that address the problem of how to achieve computational efficiency and accuracy.
Article
Analysis of the Kv3 subfamily of K(+) channel subunits has lead to the discovery of a new class of neuronal voltage-gated K(+) channels characterized by positively shifted voltage dependencies and very fast deactivation rates. These properties are adaptations that allow these channels to produce currents that can specifically enable fast repolarization of action potentials without compromising spike initiation or height. The short spike duration and the rapid deactivation of the Kv3 currents after spike repolarization maximize the quick recovery of resting conditions after an action potential. Several neurons in the mammalian CNS have incorporated into their repertoire of voltage-dependent conductances a relatively large number of Kv3 channels to enable repetitive firing at high frequencies - an ability that crucially depends on the special properties of Kv3 channels and their impact on excitability.
Article
KChIPs are a family of Kv4 K(+) channel ancillary subunits whose effects usually include slowing of inactivation, speeding of recovery from inactivation, and increasing channel surface expression. We compared the effects of the 270 amino acid KChIP2b on Kv4.3 and a Kv4.3 inner pore mutant [V(399, 401)I]. Kv4.3 showed fast inactivation with a bi-exponential time course in which the fast time constant predominated. KChIP2b expressed with wild-type Kv4.3 slowed the fast time constant of inactivation; however, the overall rate of inactivation was faster due to reduction of the contribution of the slow inactivation phase. Introduction of [V(399, 401)I] slowed both time constants of inactivation less than 2-fold. Inactivation was incomplete after 20s pulse durations. Co-expression of KChIP2b with Kv4.3 [V(399, 401)I] slowed inactivation dramatically. KChIP2b increased the rate of recovery from inactivation 7.6-fold in the wild-type channel and 5.7-fold in Kv4.3 [V(399,401)I]. These data suggest that inner pore structure is an important factor in the modulatory effects of KChIP2b on Kv4.3 K(+) channels.
Article
A rapidly inactivating K(+) current (A-type current; I(A)) present in murine colonic myocytes is important in maintaining physiological patterns of slow wave electrical activity. The kinetic profile of colonic I(A) resembles that of Kv4-derived currents. We examined the contribution of Kv4 alpha-subunits to I(A) in the murine colon using pharmacological, molecular and immunohistochemical approaches. The divalent cation Cd(2+) decreased peak I(A) and shifted the voltage dependence of activation and inactivation to more depolarized potentials. Similar results were observed with La(3+). Colonic I(A) was sensitive to low micromolar concentrations of flecainide (IC(50) = 11 microM). Quantitative PCR indicated that in colonic and jejunal tissue, Kv4.3 transcripts demonstrate greater relative abundance than transcripts encoding Kv4.1 or Kv4.2. Antibodies revealed greater Kv4.3-like immunoreactivity than Kv4.2-like immunoreactivity in colonic myocytes. Kv4-like immunoreactivity was less evident in jejunal myocytes. To address this finding, we examined the expression of K(+) channel-interacting proteins (KChIPs), which act as positive modulators of Kv4-mediated currents. Qualitative PCR identified transcripts encoding the four known members of the KChIP family in isolated colonic and jejunal myocytes. However, the relative abundance of KChIP transcript was 2.6-fold greater in colon tissue than in jejunum, as assessed by quantitative PCR, with KChIP1 showing predominance. This observation is in accordance with the amplitude of the A-type current present in these two tissues, where colonic myocytes possess densities twice that of jejunal myocytes. From this we conclude that Kv4.3, in association with KChIP1, is the major molecular determinant of I(A) in murine colonic myocytes.
Article
A low voltage-activated potassium current, IKL, is found in auditory neuron types that have low excitability and precisely preserve the temporal pattern of activity present in their presynaptic inputs. The gene Kcna1 codes for Kv1.1 potassium channel subunits, which combine in expression systems to produce channel tetramers with properties similar to those of IKL, including sensitivity to dendrotoxin (DTX). Kv1.1 is strongly expressed in neurons with IKL, including auditory neurons of the medial nucleus of the trapezoid body (MNTB). We therefore decided to investigate how the absence of Kv1.1 affected channel properties and function in MNTB neurons from mice lacking Kcna1. We used the whole cell version of the patch clamp technique to record from MNTB neurons in brainstem slices from Kcna1-null (-/-) mice and their wild-type (+/+) and heterozygous (+/-) littermates. There was an IKL in voltage-clamped -/- MNTB neurons, but it was about half the amplitude of the IKL in +/+ neurons, with otherwise similar properties. Consistent with this, -/- MNTB neurons were more excitable than their +/+ counterparts; they fired more than twice as many action potentials (APs) during current steps, and the threshold current amplitude required to generate an AP was roughly halved. +/- MNTB neurons had excitability and IKL amplitudes identical to the +/+ neurons. The IKL remaining in -/- neurons was blocked by DTX, suggesting the underlying channels contained subunits Kv1.2 and/or Kv1.6 (also DTX-sensitive). DTX increased excitability further in the already hyperexcitable -/- MNTB neurons, suggesting that -/- IKL limited excitability despite its reduced amplitude in the absence of Kv1.1 subunits.
Article
Using kinetic data from three different K+ currents in acutely isolated neurons, a single electrical compartment representing the soma of a ventral cochlear nucleus (VCN) neuron was created. The K+ currents include a fast transient current (IA), a slow-inactivating low-threshold current (ILT), and a noninactivating high-threshold current (IHT). The model also includes a fast-inactivating Na+ current, a hyperpolarization-activated cation current (Ih), and 1-50 auditory nerve synapses. With this model, the role IA, ILT, and IHT play in shaping the discharge patterns of VCN cells is explored. Simulation results indicate that IHT mainly functions to repolarize the membrane during an action potential, and IA functions to modulate the rate of repetitive firing. ILT is found to be responsible for the phasic discharge pattern observed in Type II cells (bushy cells). However, by adjusting the strength of ILT, both phasic and regular discharge patterns are observed, demonstrating that a critical level of ILT is necessary to produce the Type II response. Simulated Type II cells have a significantly faster membrane time constant in comparison to Type I cells (stellate cells) and are therefore better suited to preserve temporal information in their auditory nerve inputs by acting as precise coincidence detectors and having a short refractory period. Finally, we demonstrate that modulation of Ih, which changes the resting membrane potential, is a more effective means of modulating the activation level of ILT than simply modulating ILT itself. This result may explain why ILT and Ih are often coexpressed throughout the nervous system.
Article
Neurons in the ventral cochlear nucleus (VCN) express three distinct K+ currents that differ in their voltage and time dependence, and in their inactivation behavior. In the present study, we quantitatively analyze the voltage-dependent kinetics of these three currents to gain further insight into how they regulate the discharge patterns of VCN neurons and to provide supporting data for the identification of their channel components. We find the transient A-type K+ current (IA) exhibits fourth-order activation kinetics (a4), and inactivates with one or two time constants. A second inactivation rate (leading to an a4bc kinetic description) is required to explain its recovery from inactivation. The dendrotoxin-sensitive low-threshold K+ current (ILT) also activates with fourth-order kinetics (w4) but shows slower, incomplete inactivation. The high-threshold K+ current (IHT) appears to consist of two kinetically distinct components (n2 + p). The first component activates approximately 10 mV positive to the second and has second-order kinetics. The second component activates with first-order kinetics. These two components also contribute to two kinetically distinct currents upon deactivation. The kinetic behavior of IHT was indistinguishable amongst cell types, suggesting the current is mediated by the same K+ channels amongst VCN neurons. Together these results provide a basis for more realistic modeling of VCN neurons, and provide clues regarding the molecular basis of the three K+ currents.
Article
In the ventral cochlear nucleus (VCN), neurons transform information from auditory nerve fibers into a set of parallel ascending pathways, each emphasizing different aspects of the acoustic environment. Previous studies have shown that VCN neurons differ in their intrinsic electrical properties, including the K+ currents they express. In this study, we examine these K+ currents in more detail using whole cell voltage-clamp techniques on isolated VCN cells from adult guinea pigs at 22 degrees C. Our results show a differential expression of three distinct K+ currents. Whereas some VCN cells express only a high-threshold delayed-rectifier-like current (IHT), others express IHT in combination with a fast inactivating current (IA) and/or a slow-inactivating low-threshold current (ILT). IHT, ILT, and IA, were partially blocked by 1 mM 4-aminopyridine. In contrast, only ILT was blocked by 10-100 nM dendrotoxin-I. A surprising finding was the wide range of levels of ILT, suggesting ILT is expressed as a continuum across cell types rather than modally in a particular cell type. IA, on the other hand, appears to be expressed only in cells that show little or no ILT, the Type I cells. Boltzmann analysis shows IHT activates with 164 +/- 12 (SE) nS peak conductance, -14.3 +/- 0.7 mV half-activation, and 7.0 +/- 0.5 mV slope factor. Similar analysis shows ILT activates with 171 +/- 22 nS peak conductance, -47.4 +/- 1.0 mV half-activation, and 5.8 +/- 0.3 mV slope factor.
Article
The firing patterns of neurons in central auditory pathways encode specific features of sound stimuli, such as frequency, intensity and localization in space. The generation of the appropriate pattern depends, to a major extent, on the properties of the voltage-dependent potassium channels in these neurons. The mammalian auditory pathways that compute the direction of a sound source are located in the brainstem and include the connection from bushy cells in the anteroventral cochlear nucleus (AVCN) to the principal neurons of the medial nucleus of the trapezoid body (MNTB). To preserve the fidelity of timing of action potentials that is required for sound localization, these neurons express several types of potassium channels, including the Kv3 and Kv1 families of voltage-dependent channels and the Slick and Slack sodium-dependent channels. These channels determine the pattern of action potentials and the amount of neurotransmitter released during repeated stimulation. The amplitude of currents carried by one of these channels, the Kv3.1b channel, is regulated in the short term by protein phosphorylation, and in the long term, by changes in gene expression, such that the intrinsic excitability of the neurons is constantly being regulated by the ambient auditory environment.
Article
A molecular genetic approach was exploited to directly test the hypothesis that voltage-gated K+ (Kv) channel pore-forming (alpha) subunits of the Kv4 subfamily encode the fast transient outward K+ current (IA) in cortical pyramidal neurons and to explore the functional role of IA in shaping action potential waveforms and in controlling repetitive firing in these cells. Using the biolistic gene gun, cDNAs encoding a mutant Kv4.2 alpha subunit (Kv4.2W362F), which functions as a dominant negative (Kv4.2DN), and enhanced green fluorescent protein (EGFP) were introduced in vitro into neurons isolated from postnatal rat primary visual cortex. Whole-cell voltage-clamp recordings obtained from EGFP-positive pyramidal neurons revealed that IA is selectively eliminated in cells expressing Kv4.2DN. The densities and properties of the other Kv currents are unaffected. In neurons expressing Kv4.2DN, input resistances are increased and the (current) thresholds for action potential generation are decreased. In addition, action potential durations are prolonged, the amplitudes of afterhyperpolarizations are reduced, and the responses to prolonged depolarizing inputs are altered markedly in cells expressing Kv 4.2DN. At low stimulus intensities, firing rates are increased in Kv4.2DN-expressing cells, whereas at high stimulus intensities, Kv4.2DN-expressing cells adapt strongly. Together, these results demonstrate that Kv4alpha subunits encode IA channels and that IA plays a pivotal role in shaping the waveforms of individual action potentials and in controlling repetitive firing in visual cortical pyramidal neurons.
Article
Genes Kcna1 and Kcna2 code for the voltage-dependent potassium channel subunits Kv1.1 and Kv1.2, which are coexpressed in large axons and commonly present within the same tetramers. Both contribute to the low-voltage-activated potassium current I Kv1, which powerfully limits excitability and facilitates temporally precise transmission of information, e.g., in auditory neurons of the medial nucleus of the trapezoid body (MNTB). Kcna1-null mice lacking Kv1.1 exhibited seizure susceptibility and hyperexcitability in axons and MNTB neurons, which also had reduced I Kv1. To explore whether a lack of Kv1.2 would cause a similar phenotype, we created and characterized Kcna2-null mice (-/-). The -/- mice exhibited increased seizure susceptibility compared with their +/+ and +/- littermates, as early as P14. The mRNA for Kv1.1 and Kv1.2 increased strongly in +/+ brain stems between P7 and P14, suggesting the increasing importance of these subunits for limiting excitability. Surprisingly, MNTB neurons in brain stem slices from -/- and +/- mice were hypoexcitable despite their Kcna2 deficit, and voltage-clamped -/- MNTB neurons had enlarged I Kv1. This contrasts strikingly with the Kcna1-null MNTB phenotype. Toxin block experiments on MNTB neurons suggested Kv1.2 was present in every +/+ Kv1 channel, about 60% of +/- Kv1 channels, and no -/- Kv1 channels. Kv1 channels lacking Kv1.2 activated at abnormally negative potentials, which may explain why MNTB neurons with larger proportions of such channels had larger I Kv1. If channel voltage dependence is determined by how many Kv1.2 subunits each contains, neurons might be able to fine-tune their excitability by adjusting the Kv1.1:Kv1.2 balance rather than altering Kv1 channel density.
Article
Principal neurons of the medial nucleus of the trapezoid body (MNTB) receive a synaptic input from a single giant calyx terminal that generates a fast-rising, large excitatory postsynaptic current (EPSC), each of which are supra-threshold for postsynaptic action potential generation. Here, we present evidence that MNTB principal neurons receive multiple excitatory synaptic inputs generating slow-rising, small EPSCs that are also capable of triggering postsynaptic action potentials but are of non-calyceal origin. Both calyceal and non-calyceal EPSCs are mediated by alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) and N-methyl-d-aspartate (NMDA) receptor activation; however, the NMDA receptor-mediated response is proportionally larger at the non-calyceal synapses. Non-calyceal synapses generate action potentials in MNTB principal neurons with a longer latency and a lower reliability than the large calyceal input. They constitute an alternative low fidelity synaptic input to the fast and secure relay transmission via the calyx of Held synapse.
Article
Principal cells of the medial nucleus of the trapezoid body (MNTB) receive their excitatory input through large somatic terminals, the calyces of Held, which arise from axons of globular bushy cells located in the contralateral ventral cochlear nucleus. Discharges of MNTB neurons are characterized by high stimulus evoked firing rates, temporally precise onset responses, and a high degree of phase-locking to either pure tones or stimulus envelopes. Since the calyx of Held synapse is accessible to in vitro and to in vivo recordings, it serves as one of the most elaborate models for studying synaptic transmission in the mammalian brain. Although in such studies, the major emphasis is on synaptic physiology, the interpretation of the data will benefit from an understanding of the MNTB's contribution to auditory signal processing, including possible functional differences in different species. This implies the consideration of possible functional differences in different species. Here, we compare single unit recordings from MNTB principal cells in vivo in three different rodent species: gerbil, mouse and rat. Because of their good low-frequency hearing gerbils are often used in in vivo preparations, while mice and rats are predominantly used in slice preparations. We show that MNTB units in all three species exhibit high firing rates and precise onset-timing. Still there are species-specific specializations that might suggest the preferential use of one species over the others, depending on the scope of the respective investigation.