Afterhyperpolarization Research Articles

Modulation of BK channels contributes to activity-dependent increase of excitability through MTORC1 activity in CA1 pyramidal cells of mouse hippocampus.

Memory acquisition and synaptic plasticity are accompanied by changes in the intrinsic excitability of CA1 pyramidal neurons. These activity-dependent changes in excitability are mediated by modulation of intrinsic currents which alters the responsiveness of the cell to synaptic inputs. The afterhyperpolarization (AHP), a major contributor to the regulation of neuronal excitability, is reduced in animals that have acquired several types of hippocampus-dependent memory tasks and also following synaptic potentiation by high frequency stimulation. BK channels underlie the fast AHP and contribute to spike repolarization, and this AHP is reduced in animals that successfully acquired trace-eyeblink conditioning. This suggests that BK channel function is activity-dependent, but the mechanisms are unknown. In this study, we found that blockade of BK channels with paxilline (10 μM) decreased IAHP amplitude and increased spike half-width and instantaneous frequency in response to a +100 pA depolarization. In addition, induction of long term potentiation (LTP) by theta burst stimulation (TBS) in CA1 pyramidal neurons reduced BK channel’s contribution to IAHP, spike repolarization, and instantaneous frequency. This result indicates that BK channel activity is decreased following synaptic potentiation. Interestingly, blockade of mammalian target of rapamycin (MTORC1) with rapamycin (400 nM) following synaptic potentiation restored BK channel function, suggesting a role for protein translation in signaling events which decreased postsynaptic BK channel activity following synaptic potentiation.

Electrophysiological properties of genetically identified subtypes of layer 5 neocortical pyramidal neurons: Ca²⁺ dependence and differential modulation by norepinephrine.

We studied neocortical pyramidal neurons from two lines of bacterial artificial chromosome mice (etv1 and glt; Gene Expression Nervous System Atlas: GENSAT project), each of which expresses enhanced green fluorescent protein (EGFP) in a different subpopulation of layer 5 pyramidal neurons. In barrel cortex, etv1 and glt pyramidal cells were previously reported to differ in terms of their laminar distribution, morphology, thalamic inputs, cellular targets, and receptive field size. In this study, we measured the laminar distribution of etv1 and glt cells. On average, glt cells were located more deeply; however, the distributions of etv1 and glt cells extensively overlap in layer 5. To test whether these two cell types differed in electrophysiological properties that influence firing behavior, we prepared acute brain slices from 2-4-wk-old mice, where EGFP-positive cells in somatosensory cortex were identified under epifluorescence and then studied using whole cell current- or voltage-clamp recordings. We studied the details of action potential parameters and repetitive firing, characterized by the larger slow afterhyperpolarizations (AHPs) in etv1 neurons and larger medium AHPs (mAHPS) in glt cells, and compared currents underlying the mAHP and slow AHP (sAHP) in etv1 and glt neurons. Etv1 cells exhibited lower dV/dt for spike polarization and repolarization and reduced direct current (DC) gain (lower f-I slope) for repetitive firing than glt cells. Most importantly, we found that 1) differences in the expression of Ca(2+)-dependent K(+) conductances (small-conductance calcium-activated potassium channels and sAHP channels) determine major functional differences between etv1 and glt cells, and 2) there is differential modulation of etv1 and glt neurons by norepinephrine.

SK Channel Distribution on Neuronal Axons Revealed by Single Molecule Atomic Force Microscopy

Introduction: The axon initial segment (AIS) is a specialized cellular compartment where action potentials (APs) initiate. Calcium-activated small conductance potassium (SK) channels reduce excitability by mediating the afterhyperpolarizing (AHP) current and thus modulate the intervals between spikes during a burst of action potentials. SK channels have a polarized distribution in neurons with high density in dendrites and lower density in the soma. However, it is not clear if and which SK channels are expressed on the axon and their spatial distribution. Here, we employ single molecule atomic force microscopy (AFM) combined with natural toxins to determine the localization of SK channels on neuronal axons.Methods and Results: We have previously demonstrated that integration of single molecule AFM and toxin pharmacology allows for high resolution mapping of native SK2 channels on soma and dendrites of living neurons. AFM tip functionalized with SK channel blocker-apamin, was used to detect the distribution of SK channels by measuring the unbinding forces between apamin and SK channels. The axon was identified by transfecting rat pyramidal neurons with tau-gfp which localizes at the axon. Apamin-functionalized AFM tips were used to probe 1µm∧2 scan area on the axon of transfected pyramidal neurons. We detected unbinding forces with a mean of µ=20± 9pN and a surface density of 5.9% (n=8). To determine the specificity of the detected unbinding forces, the experiments were repeated with neurons pretreated with apamin. Along with a lower frequency of unbinding events, we found that in neurons pretreated with apamin, there was a decrease in the unbinding forces. The results above indicate that apamin sensitive SK channels reside on the axon of pyramidal neurons.Conclusion: Employing single molecule AFM, we have revealed that SK channels reside on axonal surfaces of neurons.

Electrophysiological identification of tonic and phasic neurons in sensory dorsal root ganglion and their distinct implications in inflammatory pain.

In the mammalian autonomic nervous system, tonic and phasic neurons can be differentiated on firing patterns in response to long depolarizing current pulse. However, the similar firing patterns in the somatic primary sensory neurons and their functional significance are not well investigated. Here, we identified two types of neurons innervating somatic sensory in rat dorsal root ganglia (DRG). Tonic neurons fire action potentials (APs) in an intensity-dependent manner, whereas phasic neurons typically generate only one AP firing at the onset of stimulation regardless of intensity. Combining retrograde labeling of somatic DRG neurons with fluorescent tracer DiI, we further find that these neurons demonstrate distinct changes under inflammatory pain states induced by complete Freund's adjuvant (CFA) or bee venom toxin melittin. In tonic neurons, CFA and melittin treatments significantly decrease rheobase and AP durations (depolarization and repolarization), enhance amplitudes of overshoot and afterhyperpolarization (AHP), and increase the number of evoked action potentials. In phasic neurons, however, the same inflammation treatments cause fewer changes in these electrophysiological parameters except for the increased overshoot and decreased AP durations. In the present study, we find that tonic neurons are more hyperexcitable than phasic neurons after peripheral noxious inflammatory stimulation. The results indicate the distinct contributions of two types of DRG neurons in inflammatory pain.

Morphine dependence in single enteric neurons from the mouse colon requires deletion of β-arrestin2.

Chronic administration of morphine results in the development of tolerance to the analgesic effects and to inhibition of upper gastrointestinal motility but not to colonic motility, resulting in persistent constipation. In this study we examined the effect of chronic morphine in myenteric neurons from the adult mouse colon. Similar to the ileum, distinct neuronal populations exhibiting afterhyperpolarization (AHP)‐positive and AHP‐negative neurons were identified in the colon. Acute morphine (3 μM) decreased the number of action potentials, and increased the threshold for action potential generation indicative of reduced excitability in AHP‐positive neurons. In neurons from the ileum of mice that were rendered antinociceptive tolerant by morphine‐pellet implantation for 5 days, the opioid antagonist naloxone precipitated withdrawal as evidenced by increased neuronal excitability. Overnight incubation of ileum neurons with morphine also resulted in enhanced excitability to naloxone. Colonic neurons exposed to long‐term morphine, remained unresponsive to naloxone suggesting that precipitated withdrawal does not occur in colonic neurons. However, morphine‐treated colonic neurons from β‐arrestin2 knockout mice demonstrated increased excitability upon treatment with naloxone as assessed by change in rheobase, number of action potentials and input resistance. These data suggest that similar to the ileum, acute exposure to morphine in colonic neurons results in reduced excitability due to inhibition of sodium currents. However, unlike the ileum, dependence to chronic exposure of morphine develops in colonic neurons from the β‐arrestin2 knockout mice. These studies corroborate the in‐vivo findings of the differential role of neuronal β‐arrestin2 in the development of morphine tolerance/dependence in the ileum and colon.

Decreased calcium-activated potassium channels by hypoxia causes abnormal firing in the spontaneous firing medial vestibular nuclei neurons.

Vertebrobasilar insufficiency (VBI) presents complex varied clinical symptoms, including vertigo and hearing loss. Little is known, however, about how Ca(2+)-activated K(+) channel attributes to the medial vestibular nucleus (MVN) neural activity in VBI. To address this issue, we performed whole-cell patch clamp and quantitative polymerase chain reaction (qPCR) to examine the effects of hypoxia on neural activity and the changes of the large conductance Ca(2+) activated K(+) channels (BKCa channels) in the MVN neurons in brain slices of male C57BL/6 mice. Brief hypoxic stimuli of the brain slices containing MVN were administrated by switching the normoxic artificial cerebrospinal fluid (ACSF) equilibrated with 21% O2/5% CO2 to hypoxic ACSF equilibrated with 5% O2/5% CO2 (balance N2). 3-min hypoxia caused a depolarization in the resting membrane potential (RM) in 8/11 non-spontaneous firing MVN neurons. 60/72 spontaneous firing MVN neurons showed a dramatic increase in firing frequency and a depolarization in the RM following brief hypoxia. The amplitude of the afterhyperpolarization (AHPA) was significantly decreased in both type A and type B spontaneous firing MVN neurons. Hypoxia-induced firing response was alleviated by pretreatment with NS1619, a selective BKCa activator. Furthermore, brief hypoxia caused a decrease in the amplitude of iberiotoxin-sensitive outward currents and mRNA level of BKCa in MVN neurons. These results suggest that BKCa channels protect against abnormal MVN neuronal activity induced by hypoxia, and might be a key target for treatment of vertigo and hearing loss in VBI.

Increased intrinsic excitability of muscle vasoconstrictor preganglionic neurons may contribute to the elevated sympathetic activity in hypertensive rats.

Hypertension is associated with pathologically increased sympathetic drive to the vasculature. This has been attributed to increased excitatory drive to sympathetic preganglionic neurons (SPN) from brainstem cardiovascular control centers. However, there is also evidence supporting increased intrinsic excitability of SPN. To test this hypothesis, we made whole cell recordings of muscle vasoconstrictor-like (MVClike) SPN in the working-heart brainstem preparation of spontaneously hypertensive (SH) and normotensive Wistar-Kyoto (WKY) rats. The MVClike SPN have a higher spontaneous firing frequency in the SH rat (3.85 ± 0.4 vs. 2.44 ± 0.4 Hz in WKY; P = 0.011) with greater respiratory modulation of their activity. The action potentials of SH SPN had smaller, shorter afterhyperpolarizations (AHPs) and showed diminished transient rectification indicating suppression of an A-type potassium conductance (IA). We developed mathematical models of the SPN to establish if changes in their intrinsic properties in SH rats could account for their altered firing. Reduction of the maximal conductance density of IA by 15–30% changed the excitability and output of the model from the WKY to a SH profile, with increased firing frequency, amplified respiratory modulation, and smaller AHPs. This change in output is predominantly a consequence of altered synaptic integration. Consistent with these in silico predictions, we found that intrathecal 4-aminopyridine (4-AP) increased sympathetic nerve activity, elevated perfusion pressure, and augmented Traube-Hering waves. Our findings indicate that IA acts as a powerful filter on incoming synaptic drive to SPN and that its diminution in the SH rat is potentially sufficient to account for the increased sympathetic output underlying hypertension.

Context-dependent coding in single neurons.

The linear-nonlinear cascade model (LN model) has proven very useful in representing a neural system's encoding properties, but has proven less successful in reproducing the firing patterns of individual neurons whose behavior is strongly dependent on prior firing history. While the cell's behavior can still usefully be considered as feature detection acting on a fluctuating input, some of the coding capacity of the cell is taken up by the increased firing rate due to a constant "driving" direct current (DC) stimulus. Furthermore, both the DC input and the post-spike refractory period generate regular firing, reducing the spike-timing entropy available for encoding time-varying fluctuations. In this paper, we address these issues, focusing on the example of motoneurons in which an afterhyperpolarization (AHP) current plays a dominant role regularizing firing behavior. We explore the accuracy and generalizability of several alternative models for single neurons under changes in DC and variance of the stimulus input. We use a motoneuron simulation to compare coding models in neurons with and without the AHP current. Finally, we quantify the tradeoff between instantaneously encoding information about fluctuations and about the DC.

Afferent-hair cell connectivity as a possible source of spike train irregularity in turtle vestibular bouton afferents

Vestibular afferents have been categorized as “regular” or “irregular” depending on the variability of their spike trains [1,2]. It is thought that the difference in regularity among afferents results from differences in the types of voltage-dependent conductances present in the cell. For example afferents may differ in their complements of potassium A, D, and M currents or in currents that produce a small vs. prominent afterhyperpolarization (AHP) [3-5]. Because bouton afferents from turtle utricle vary widely in morphology depending on their location in the utricle, we sought to determine if afferent morphology might also play a role in spike train irregularity. Although we found that morphology itself is not a factor in determining spike train variability, we did find that spike train variability can be affected by afferent/hair cell connectivity. Morphological reconstructions of bouton afferents from the lateral extrastriolar (LES), striola, juxtastriola and medial extrastriolar (MES) regions of turtle utricle were done with Neurolucida. Models based on these reconstructions were constructed for 16 afferents. Voltage-dependent conductance distributions in different regions (axon, soma, terminal arbor, varicosities, nodes, myelin) were identical in each cell model. Given the locations of synapses in the terminal arbor and the density of hair cells in each region, we were able to estimate the number of synapses formed between individual hair cells and an afferent. In the LES, striola and juxtastriola most synapses on an afferent were formed with unique hair cells, whereas in the MES single hair cells more often made multiple synapses with an afferent. To determine the effect of this difference in connectivity we modeled Poisson release from hair cells to synapses in the terminal arbor to generate spike trains. The coefficient of variation (cv) as a function of interspike interval (ISI) was calculated to estimate spike train variability with different average release rates. We found that cv as a function of ISI was nearly identical for LES, striola and juxtastriola afferents, but was the same for MES afferents only if connectivity in the MES was assumed to be one afferent synapse per hair cell. However, cv was much larger when the MES afferents were modeled with multiple synapses between an afferent and a hair cell, as calculated from synaptic location and hair cell density. To see if we could make an LES afferent more irregular, we modified connectivity to have more multiple synapses between individual hair cells and the afferent. As predicted, cv increased. These results suggest that afferent-hair cell connectivity may be a source of spike train irregularity. Given the large number of multiple contacts between a calyx and its presynaptic hair cell, we expect that connectivity contributes to spike train variability in calyx-bearing (calyx and dimorph) afferents.

Maturation of membrane properties of neurons in the rat deep cerebellar nuclei.

Patch clamp recordings of neurons in the adult rat deep cerebellar nuclei have been limited by the availability of viable brain slices. Using a new slicing technique, this study was designed to explore the maturation of membrane properties of neurons in the deep cerebellar nuclei (DCN)-an area involved in rat eyeblink conditioning. Compared to whole-cell current-clamp recordings in DCN in rat pups at postnatal day 16 (P16) to P21, recordings from weanling rats at P22-P40 revealed a number of significant changes including an increase in the amplitude of the afterhyperpolarization (AHP)-an index of membrane excitability which has been shown to be important for eyeblink conditioning-a prolonged interval between the first and second evoked action potential, and an increase in AHP amplitude for hyperpolarization-induced rebound spikes. This is the first report of developmental changes in membrane properties of DCN which may contribute to the ontogeny of eyeblink conditioning in the rat.

Changes in motoneuron afterhyperpolarization duration in stroke survivors.

Hemispheric brain injury resulting from a stroke is often accompanied by muscle weakness in limbs contralateral to the lesion. In the present study, we investigated whether weakness in contralesional hand muscle in stroke survivors is partially attributable to alterations in motor unit activation, including alterations in firing rate modulation range. The afterhyperpolarization (AHP) potential of a motoneuron is a primary determinant of motoneuron firing rate. We examined differences in AHP duration in motoneurons innervating paretic and less impaired (contralateral) limb muscles of hemiparetic stroke survivors as well as in control subjects. A novel surface EMG (sEMG) electrode was used to record motor units from the first dorsal interosseous muscle. The sEMG data were subsequently decomposed to derive single-motor unit events, which were then utilized to produce interval (ISI) histograms of the motoneuron discharges. A modified version of interval death rate (IDR) analysis was used to estimate AHP duration. Results from data analyses performed on both arms of 11 stroke subjects and in 7 age-matched control subjects suggest that AHP duration is significantly longer for motor units innervating paretic muscle compared with units in contralateral muscles and in units of intact subjects. These results were supported by a coefficient of variation (CV) analysis showing that paretic motor unit discharges have a lower CV than either contralateral or control units. This study suggests that after stroke biophysical changes occur at the motoneuron level, potentially contributing to lower firing rates and potentially leading to less efficient force production in paretic muscles.

Apamin-sensitive potassium current and the Mechanisms of ventricular arrhythmia: Perspectives in heart failure treatment

In patients with advanced heart failure (HF), recurrent spontaneous ventricular fibrillation (SVF)/electrical storm (ES) is a frequent complication requiring multiple defibrillation shocks within a short space of time. The mechanism underlying initiation of SVF/ES, spontaneously or immediately after defibrillation shocks, remains unclear. In 2009, Ogawa et al. developed a pacing-induced HF model, in which SVF occurred frequently in failing rabbit ventricles [1]. In that model, acute but reversible post-shock action potential duration (APD) shortening could induce recurrent SVF (Fig. 1, panel A) [1]. The induction of SVF was associated with persistent intracellular calcium (Cai) elevation during late phase 3 and/or phase 4 of the action potential (AP), indicating that APD shortening, in conjunction with persistent post-shock Cai elevation, is a novel mechanism of post-shock SVF/ ES [1]. However, the mechanisms underlying acute APD shortening after fibrillation/defibrillation episodes in HF ventricles remains to be determined. HF is associated with the down-regulation of multiple potassium (K) currents (i.e., Ito, IKs, IKr, IK1, and IKATP). The only K channel that is upregulated in HF is the small conductance Ca-activated K (SK) channel. The SK channel was originally found in most neurons. The increase in Cai evoked by the AP allows the SK channel to become activated, and generates a long-lasting afterhyperpolarization (AHP). Physiologically, the AHP can inhibit repetitive firing, and prevent deleterious tetanic activity in the nervous system. Previous studies show that the SK current is abundantly present in cardiac atrial cells, but not in normal ventricular cells. Because ventricular fibrillation (VF) is associated with Cai accumulation, especially in HF ventricles, Chua et al. hypothesized that SK channels might exist in HF ventricles and contribute to the post-shock APD shortening observed in Ogawa's study [1,2]. To test this hypothesis, they used apamin, a selective SK channel blocker that specifically inhibits apamin-sensitive K current (IKAS), to explore the roles of the SK channel in mediating ventricular arrhythmia of failing ventricles. Chua et al. found that apamin administration effectively prevented post-shock APD shortening, late phase 3 early after-depolarizations (EAD), and triggered activity and recurrent SVF in failing rabbit ventricles (see Fig. 1, panel B) [2]. Using a voltage-clamp technique, they reported that the IKAS current density was significantly larger in failing ventricular cardiomyocytes than in normal ones [2]. The IKAS sensitivity to Cai was increased in cardiomyocytes isolated from failing ventricles compared to those from normal ventricles [2]. Similar findings were reproduced in failing human ventricles by Chang et al. [3]. They also showed that the IKAS current density was lower in the mid-myocardial cells than in the epicardial and endocardial cells [3]. The subtype 2 of SK (SK2) protein expression was 3-fold higher in HF than in non-HF human ventricles [3]. These findings indicated for the first time that HF heterogeneously increases the sensitivity of IKAS to Cai, leading to the up-regulation of IKAS, post-shock APD shortening, late phase 3 EAD, triggered activity, and recurrent SVF in both animal models and failing human ventricles.

Afterhyperpolarization of human motoneurons firing double and triple discharges.

During isometric voluntary contractions of a healthy human muscle, motoneurons (MNs) fire usually with low mean rates, rarely exceeding 25/s (e.g., Garland and Griffin, 1999). However, there are MNs, which sometimes fire double discharges (doublets) with interspike interval (ISI) of few ms. This is observed seldom in normal MNs (e.g., Denslow, 1948; Kudina, 1974; Bawa and Calancie, 1983) but does so more often in neuromuscular disorders (Partanen, 1978; Kostera-Pruszczyk et al., 2002; Piotrkiewicz et al., 2008), where it is considered to be an early sign of MN dysfunction (Rowinska-Marcinska et al., 1999). Our special attention is paid to the so-called “true” doublets recorded in some MNs under conditions of constant synaptic drive, as opposed to those recorded during rapid contractions (Bawa and Calancie, 1983; Kudina and Andreeva, 2013a). Initial doublets, observed during repetitive movements such as locomotion (Zajac and Young, 1980; Hennig and Lomo, 1987) or respiration (Kirkwood and Munson, 1996), where the synaptic drive undergoes periodic changes, also cannot be classified as “true” doublets. For the sake of clarity, the adjective “true” will be omitted in the further text. It has been observed that not each human MN is capable of firing doublets (Kudina, 1974; Bawa and Calancie, 1983). By the analogy to animal studies, it was hypothesized that doublets may be generated only in these MNs, which possess the delayed depolarization (DD, Granit et al., 1963; Kernell, 1964; Calvin, 1974) with a prominent hump that may spontaneously cross the firing threshold and evoke an extra spike (Nelson and Burke, 1967; Calvin, 1973). This hypothesis was verified by Kudina and Churikova (1990), who tested the changes in excitability of human motoneurons within ISI by studying the responses of single MNs to the stimulation of Ia afferents. In MNs capable of firing doublets they revealed the period of increased excitability during the first 15 ms after regular discharge, which corresponded well to the duration of DD observed in animal experiments (Granit et al., 1963; Kernell, 1964; Nelson and Burke, 1967; Calvin, 1973). Nowadays, DD is widely accepted as the mechanism responsible for doublet generation in human MNs (e.g., Kudina, 1974; Bawa and Calancie, 1983; Kudina and Churikova, 1990; Garland and Griffin, 1999). However, it can explain only the origin of occasional doublets, interspersed in regular rhythmic MN activity. Repetitive doublets, i.e., series of doublet-postdoublet ISIs, require some additional mechanism, which would support the DD hump, shown to disappear during MN rhythmic firing (Granit et al., 1963; Calvin and Loeser, 1975). It was recently hypothesized that repetitive doublet firing may be related to plateau potentials (Kudina and Andreeva, 2010, 2013a). It is still under debate, whether DD is related to other characteristics of a MN, measurable in human experiments. Kernell (1964) noted that DD with a definite hump had a tendency to be more common in MNs with short afterhyperpolarization (AHP), although it could occur with an AHP of any duration. Also in humans, MNs capable of firing doublets were more often found in the fast muscles (Bawa and Calancie, 1983; Kudina and Alexeeva, 1992) than in the slower ones (Andreassen and Rosenfalck, 1980; Kudina and Andreeva, 2013b). For the long time it was believed that the MNs supplying one of the most often investigated slow human muscle, the soleus, are devoid of DD. However, it has been recently shown that also MNs supplying human soleus may fire doublets (Piotrkiewicz et al., 2013), which seems to question Kernell's finding. In the present paper we will compare the estimates of AHP duration of MNs capable and not capable of firing double discharges and comment on this controversy. We will also propose a hypothesis on the AHP-related mechanism underlying firing of triplets, which can be observed in some doublet firing MNs.

D-Cycloserine enhances both intrinsic excitability of CA1 hippocampal neurons and expression of activity-regulated cytoskeletal (Arc) protein

The interaction of NMDA receptor (NMDAR) activation and other mechanisms regulating neuronal excitability have not been thoroughly described. While excess activation of NMDARs results in excitotoxicity, partial activation of NMDARs by d-cycloserine (DCS) is nootropic, enhancing both acquisition and extinction of memories. The mechanism by which DCS treatment enhances memory is unclear. NMDAR activation has been shown to increase expression of the activity-regulated cytoskeletal (Arc) protein associated with neural plasticity and enhanced memory. Enhanced memory is also associated with increases in neuronal intrinsic excitability, i.e. reductions in post-burst afterhyperpolarizations (AHPs) after acquisition of new tasks. Reductions in AHPs can occur when Ca2+ influx is reduced. This study aimed to determine if either if Arc expression, intrinsic excitability, or both were altered following systemic administration of a memory-enhancing dose of DCS, i.e. what form of plasticity would be exhibited. Both Arc protein expression and intrinsic excitability were enhanced in tissue prepared 1h post-administration of a nootropic dose of DCS. Both mechanisms have been strongly associated with memory enhancement, but have not previously been demonstrated to change across the same time frame in the same preparation in response to DCS treatment.

Computational modeling of the effects of temperature on chemosensitive locus coeruleus neurons from bullfrogs (1128.3)

Chemosensitive LC neurons from bullfrogs increase their firing rate and have altered action potential (AP) waveforms upon cooling from 20°C to 10°C (Santin et al., Amer. J. Physiol. Doi:10.1152/ajpregu.00348.2013). We developed a computational model to study this unusual neuronal response to cold temperature. Using MatLab, we have a 4 dimensional model based on a Hodgkin‐Huxley formulation that includes a voltage‐gated Na channel, a delayed‐rectifying K (Kdr) channel and a leak K channel. The model generates a firing rate of ~12 Hz and AP waveforms that are similar to those of bullfrog LC neurons at 20°C. For instance, values (neuron:model) for AP height (60 mV:51 mV), maximum rate of depolarization (36 mV/ms:40 mV/ms), maximum rate of repolarization (‐27 mV/ms:‐23 mV/ms), the AP width (2.1 ms:2.6 ms) and after hyperpolarization (AHP) (‐60 mV:‐66 mV) are similar comparing the model to the actual neuron. Colder temperature (10°C) decreases neuronal maximum rates of depolarization and repolarization, increases the AP width, and decreases the AHP. Changing, in a depolarizing direction, the voltage at which the time constant of inactivation for Kdr channels reaches a maximum and increasing the time constant for Na channel activation in the model results in similar changes in AP waveform parameters. However, the model changes result in decreased firing rate (~6 Hz) while temperature changes in neurons result in increased firing rate. This suggests that another channel is involved in the temperature effects on firing rate. In line with this, we found a hyperpolarization‐activated current (IH) in LC neurons that will be added in our model to determine if altering the parameters of the IH current contributes to cold temperature‐induced increased firing rate in bullfrog LC neurons. We ultimately plan to use both the model and electrophysiology studies to gain insights into the neuronal basis for chemosensitivity and cold temperature responses in chemosensitive LC neurons from bullfrog.Grant Funding Source: Supported by Admin. Dept. of Sci., Technol. & Innov. of Col (COLCIENCIAS) and NSF Grant IOS‐1257338.

Simvastatin Treatment Enhances NMDAR-Mediated Synaptic Transmission by Upregulating the Surface Distribution of the GluN2B Subunit

The ramifications of statins on plasma cholesterol and coronary heart disease have been well documented. However, there is increasing evidence that inhibition of the mevalonate pathway may provide independent neuroprotective and procognitive pleiotropic effects, most likely via inhibition of isoprenoids, mainly farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP). FPP and GGPP are the major donors of prenyl groups for protein prenylation. Modulation of isoprenoid availability impacts a slew of cellular processes including synaptic plasticity in the hippocampus. Our previous work has demonstrated that simvastatin (SV) administration improves hippocampus-dependent spatial memory, rescuing memory deficits in a mouse model of Alzheimer's disease. Treatment of hippocampal slices with SV enhances long-term potentiation (LTP), and this effect is dependent on the activation of Akt (protein kinase B). Further studies showed that SV-induced enhancement of hippocampal LTP is driven by depletion of FPP and inhibition of farnesylation. In the present study, we report the functional consequences of exposure to SV at cellular/synaptic and molecular levels. While application of SV has no effect on intrinsic membrane properties of CA1 pyramidal neurons, including hyperpolarization-activated cyclic-nucleotide channel-mediated sag potentials, the afterhyperpolarization (AHP), and excitability, SV application potentiates the N-methyl D-aspartate receptor (NMDAR)-mediated contribution to synaptic transmission. In mouse hippocampal slices and human neuronal cells, SV treatment increases the surface distribution of the GluN2B subunit of the NMDAR without affecting cellular cholesterol content. We conclude that SV-induced enhancement of synaptic plasticity in the hippocampus is likely mediated by augmentation of synaptic NMDAR components that are largely responsible for driving synaptic plasticity in the CA1 region.

Intracellular Gold Nanoparticles Increase Neuronal Excitability and Aggravate Seizure Activity in the Mouse Brain

Due to their inert property, gold nanoparticles (AuNPs) have drawn considerable attention; their biological application has recently expanded to include nanomedicine and neuroscience. However, the effect of AuNPs on the bioelectrical properties of a single neuron remains unknown. Here we present the effect of AuNPs on a single neuron under physiological and pathological conditions in vitro. AuNPs were intracellularly applied to hippocampal CA1 neurons from the mouse brain. The electrophysiological property of CA1 neurons treated with 5- or 40-nm AuNPs was assessed using the whole-cell patch-clamp technique. Intracellular application of AuNPs increased both the number of action potentials (APs) and input resistance. The threshold and duration of APs and the after hyperpolarization (AHP) were decreased by the intracellular AuNPs. In addition, intracellular AuNPs elicited paroxysmal depolarizing shift-like firing patterns during sustained repetitive firings (SRF) induced by prolonged depolarization (10 sec). Furthermore, low Mg2+-induced epileptiform activity was aggravated by the intracellular AuNPs. In this study, we demonstrated that intracellular AuNPs alter the intrinsic properties of neurons toward increasing their excitability, and may have deleterious effects on neurons under pathological conditions, such as seizure. These results provide some considerable direction on application of AuNPs into central nervous system (CNS).

Distinct Effects of Repeated Restraint Stress on Basolateral Amygdala Neuronal Membrane Properties in Resilient Adolescent and Adult Rats

Severe and repeated stress has damaging effects on health, including initiation of depression and anxiety. Stress that occurs during development has long-lasting and particularly damaging effects on emotion. The basolateral amygdala (BLA) plays a key role in many affective behaviors, and repeated stress causes different forms of BLA hyperactivity in adolescent and adult rats. However, the mechanism is not known. Furthermore, not every individual is susceptible to the negative consequences of stress. Differences in the effects of stress on the BLA might contribute to determine whether an individual will be vulnerable or resilient to the effects of stress on emotion. The purpose of this study is to test the cellular underpinnings for age dependency of BLA hyperactivity after stress, and whether protective changes occur in resilient individuals. To test this, the effects of repeated stress on membrane excitability and other membrane properties of BLA principal neurons were compared between adult and adolescent rats, and between vulnerable and resilient rats, using in vitro whole-cell recordings. Vulnerability was defined by adrenal gland weight, and verified by body weight gain after repeated restraint stress, and fecal pellet production during repeated restraint sessions. We found that repeated stress increased the excitability of BLA neurons, but in a manner that depended on age and BLA subnucleus. Furthermore, stress resilience was associated with an opposite pattern of change, with increased slow afterhyperpolarization (AHP) potential, whereas vulnerability was associated with decreased medium AHP. The opposite outcomes in these two populations were further distinguished by differences of anxiety-like behavior in the elevated plus maze that were correlated with BLA neuronal excitability and AHP. These results demonstrate a substrate for BLA hyperactivity after repeated stress, with distinct membrane properties to target, as well as age-dependent factors that contribute to resilience to the effects of stress.

Maternal mobile phone exposure alters intrinsic electrophysiological properties of CA1 pyramidal neurons in rat offspring

Some studies have shown that exposure to electromagnetic field (EMF) may result in structural damage to neurons. In this study, we have elucidated the alteration in the hippocampal function of offspring Wistar rats (n = 8 rats in each group) that were chronically exposed to mobile phones during their gestational period by applying behavioral, histological, and electrophysiological tests. Rats in the EMF group were exposed to 900 MHz pulsed-EMF irradiation for 6 h/day. Whole cell recordings in hippocampal pyramidal cells in the mobile phone groups did show a decrease in neuronal excitability. Mobile phone exposure was mostly associated with a decrease in the number of action potentials fired in spontaneous activity and in response to current injection in both male and female groups. There was an increase in the amplitude of the afterhyperpolarization (AHP) in mobile phone rats compared with the control. The results of the passive avoidance and Morris water maze assessment of learning and memory performance showed that phone exposure significantly altered learning acquisition and memory retention in male and female rats compared with the control rats. Light microscopy study of brain sections of the control and mobile phone-exposed rats showed normal morphology.Our results suggest that exposure to mobile phones adversely affects the cognitive performance of both female and male offspring rats using behavioral and electrophysiological techniques.

The vestibulo‐ and preposito‐cerebellar cholinergic neurons of a ChAT‐tdTomato transgenic rat exhibit heterogeneous firing properties and the expression of various neurotransmitter receptors

Cerebellar function is regulated by cholinergic mossy fiber inputs that are primarily derived from the medial vestibular nucleus (MVN) and prepositus hypoglossi nucleus (PHN). In contrast to the growing evidence surrounding cholinergic transmission and its functional significance in the cerebellum, the intrinsic and synaptic properties of cholinergic projection neurons (ChPNs) have not been clarified. In this study, we generated choline acetyltransferase (ChAT)-tdTomato transgenic rats, which specifically express the fluorescent protein tdTomato in cholinergic neurons, and used them to investigate the response properties of ChPNs identified via retrograde labeling using whole-cell recordings in brainstem slices. In response to current pulses, ChPNs exhibited two afterhyperpolarisation (AHP) profiles and three firing patterns; the predominant AHP and firing properties differed between the MVN and PHN. Morphologically, the ChPNs were separated into two types based on their soma size and dendritic extensions. Analyses of the firing responses to time-varying sinusoidal current stimuli revealed that ChPNs exhibited different firing modes depending on the input frequencies. The maximum frequencies in which each firing mode was observed were different between the neurons that exhibited distinct firing patterns. Analyses of the current responses to the application of neurotransmitter receptor agonists revealed that the ChPNs expressed (i) AMPA- and NMDA-type glutamate receptors, (ii) GABAA and glycine receptors, and (iii) muscarinic and nicotinic acetylcholine receptors. The current responses mediated by these receptors of MVN ChPNs were not different from those of PHN ChPNs. These findings suggest that ChPNs receive various synaptic inputs and encode those inputs appropriately across different frequencies.