Channels In Pyramidal Neurons Research Articles

Distinct modulation of Ih by synaptic potentiation in excitatory and inhibitory neurons.

Selective modifications in the expression or function of dendritic ion channels regulate the propagation of synaptic inputs and determine the intrinsic excitability of a neuron. Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels open upon membrane hyperpolarization and conduct a depolarizing inward current (Ih). HCN channels are enriched in the dendrites of hippocampal pyramidal neurons where they regulate the integration of synaptic inputs. Synaptic plasticity can bidirectionally modify dendritic HCN channels in excitatory neurons depending on the strength of synaptic potentiation. In inhibitory neurons, however, the dendritic expression and modulation of HCN channels is largely unknown. In this study, we systematically compared the modulation of Ih by synaptic potentiation in hippocampal CA1 pyramidal neurons and stratum Radiatum (sRad) interneurons in mouse organotypic cultures. Ih properties were similar in inhibitory and excitatory neurons and contributed to resting membrane potential and action potential firing. We found that in sRad interneurons, HCN channels were downregulated after synaptic plasticity, irrespective of the strength of synaptic potentiation. This suggest differential regulation of Ih in excitatory and inhibitory neurons, possibly signifying their distinct role in network activity.Significance statement Learning reflects a change in the way information is processed in neuronal circuits. This occurs via changes in synaptic connections and via alterations of intrinsic excitability of neurons. Here we examined how synaptic changes affect properties of HCN channels, which are important ion channels for intrinsic excitability. We found that strong synaptic potentiation leads to opposite changes in HCN channels in CA1 pyramidal neurons and sRad interneurons. We speculate that this reflects their differential role in the CA1 network. An upregulation of HCN channels in pyramidal neurons results in a decrease in their excitability, which limits overall network excitation. In contrast, sRad interneurons show downregulation of Ih, and therefore an increased excitability after strong synaptic activation, which will strengthen feedforward inhibition and sharpen activity patterns.

D-serine reconstitutes synaptic and intrinsic inhibitory control of pyramidal neurons in a neurodevelopmental mouse model for schizophrenia

The hypothesis of N-methyl-D-aspartate receptor (NMDAR) dysfunction for cognitive impairment in schizophrenia constitutes the theoretical basis for the translational application of NMDAR co-agonist D-serine or its analogs. However, the cellular mechanism underlying the therapeutic effect of D-serine remains unclear. In this study, we utilize a mouse neurodevelopmental model for schizophrenia that mimics prenatal pathogenesis and exhibits hypoexcitability of parvalbumin-positive (PV) neurons, as well as PV-preferential NMDAR dysfunction. We find that D-serine restores excitation/inhibition balance by reconstituting both synaptic and intrinsic inhibitory control of cingulate pyramidal neurons through facilitating PV excitability and activating small-conductance Ca2+-activated K+ (SK) channels in pyramidal neurons, respectively. Either amplifying inhibitory drive via directly strengthening PV neuron activity or inhibiting pyramidal excitability via activating SK channels is sufficient to improve cognitive function in this model. These findings unveil a dual mechanism for how D-serine improves cognitive function in this model.

Improved HCN channels in pyramidal neurons and their new expression levels in pericytes and astrocytes in the gerbil hippocampal CA1 subfield following transient ischemia.

Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels have been known to participate in the regulation of neuronal excitability, synaptic transmission and long-term potentiation in the hippocampus. The present study investigated transient ischemia-induced changes of HCN1 and HCN2 expressions in the Cornu Ammonis 1 (CA1) subfield of the hippocampus in gerbils subjected to 5 min transient global cerebral ischemia (tgCI). Neuronal death was exhibited in pyramidal neurons of the striatum pyramidale in the CA1 subfield 4 days after tgCI. HCN1 and HCN2 immunoreactivities were demonstrated in intact CA1 pyramidal neurons, and were transiently and markedly increased in the CA pyramidal neurons at 6 h after ischemia. Thereafter, they gradually decreased in a time-dependent manner. A total of 4 days after ischemia, HCN1 and HCN2 immunoreactivities were barely detected in the CA1 pyramidal neurons; however, HCN1 and HCN2 were began to be expressed in pericytes and astrocytes at 4 days after ischemia. The results indicated that HCN1 and HCN2 expression levels were apparently changed in the gerbil hippocampal CA1 subfield following tgCI and suggested that ischemia-induced alterations in HCN1 and HCN2 expression levels may be closely associated with the death of CA1 pyramidal neurons following 5 min of tgCI.

MGluR5-dependent modulation of dendritic excitability in CA1 pyramidal neurons mediated by enhancement of persistent Na+ currents.

High-frequency stimulation (HFS) of the Schaffer collateral pathway activates metabotropic glutamate receptor 5 (mGluR5) signalling in the proximal apical dendrites of CA1 pyramidal neurons. The synaptic activation of mGluR5-mediated calcium signalling causes a significant increase in persistent sodium current (INa,P ) in the dendrites. Increased INa,P by HFS underlies potentiation of synaptic inputs at both the proximal and distal dendrite, leading to an enhanced probability of action potential firing associated with decreased action potential thresholds. Therefore, HFS-induced activation of intracellular mGluR5 serves an important role as an instructive signal for potentiation of upcoming inputs by increasing dendritic excitability. Dendritic Na+ channels in pyramidal neurons are known to amplify synaptic signals, thereby facilitating action potential (AP) generation. However, the mechanisms that modulate dendritic Na+ channels have remained largely uncharacterized. Here, we report a new form of short-term plasticity in which proximal excitatory synaptic inputs to hippocampal CA1 pyramidal neurons transiently elevate dendritic excitability. High-frequency stimulations (HFS) to the Schaffer collateral (SC) pathway activate mGluR5-dependent Ca2+ signalling in the apical dendrites, which, with calmodulin, upregulates specifically Nav1.6 channel-mediated persistent Na+ currents (INa,P ) in the dendrites. This HFS-induced increase in dendritic INa,P results in transient increases in the amplitude of excitatory postsynaptic potentials induced by both proximal SC and distal perforant path stimulation, leading to the enhanced probability of AP firing associated with decreased AP thresholds. Taken together, our study identifies dendritic INa,P as a novel target for mediating activity-dependent modulation of dendritic integration and neuronal output.

Cell-Type Specific Development of the Hyperpolarization-Activated Current, Ih, in Prefrontal Cortical Neurons.

H-current, also known as hyperpolarization-activated current (Ih), is an inward current generated by the hyperpolarization-activated cyclic nucleotide-gated (HCN) cation channels. Ih plays an essential role in regulating neuronal properties, synaptic integration and plasticity, and synchronous activity in the brain. As these biological factors change across development, the brain undergoes varying levels of vulnerability to disorders like schizophrenia that disrupt prefrontal cortex (PFC)-dependent function. However, developmental changes in Ih in PFC neurons remains untested. Here, we examine Ih in pyramidal neurons vs. gamma-aminobutyric acid (GABA)ergic parvalbumin-expressing (PV+) interneurons in developing mouse PFC. Our findings show that the amplitudes of Ih in these cell types are identical during the juvenile period but differ at later time points. In pyramidal neurons, Ih amplitude significantly increases from juvenile to adolescence and follows a similar trend into adulthood. In contrast, the amplitude of Ih in PV+ interneurons decreases from juvenile to adolescence, and does not change from adolescence to adulthood. Moreover, the kinetics of HCN channels in pyramidal neurons is significantly slower than in PV+ interneurons, with a gradual decrease in pyramidal neurons and a gradual increase in PV+ cells across development. Our study reveals distinct developmental trajectories of Ih in pyramidal neurons and PV+ interneurons. The cell-type specific alteration of Ih during the critical period from juvenile to adolescence reflects the contribution of Ih to the maturation of the PFC and PFC-dependent function. These findings are essential for a better understanding of normal PFC function, and for elucidating Ih’s crucial role in the pathophysiology of neurodevelopmental disorders.

Suppression of inhibitory G protein signaling in forebrain pyramidal neurons triggers plasticity of glutamatergic neurotransmission in the nucleus accumbens core

Cocaine and other drugs of abuse trigger long-lasting adaptations in excitatory and inhibitory neurotransmission in the mesocorticolimbic system, and this plasticity has been implicated in several key facets of drug addiction. For example, glutamatergic neurotransmission mediated by AMPA receptors (AMPAR) is strengthened in medium spiny neurons (MSNs) in the NAc core and shell during withdrawal following repeated in vivo cocaine administration. Repeated cocaine administration also suppresses inhibitory signaling mediated by G protein-gated inwardly rectifying K+ (GIRK) channels in pyramidal neurons of the prelimbic cortex, an important source of glutamatergic input to the NAc core that has been implicated in cocaine-seeking and behavioral sensitization. Here, we tested the hypothesis that suppression of GIRK channel activity in forebrain pyramidal neurons can promote plasticity of glutamatergic signaling in MSNs. Using novel conditional knockout mouse lines, we report that GIRK channel ablation in forebrain pyramidal neurons is sufficient to enhance AMPAR-dependent neurotransmission in D1R-expressing MSNs in the NAc core, while also increasing motor-stimulatory responses to cocaine administration. A similar increase in AMPAR-dependent signaling was seen in both D1R- and D2R-expressing MSNs in the NAc core during withdrawal from repeated cocaine administration in normal mice. Collectively, these data are consistent with the premise that the cocaine-induced suppression of GIRK-dependent signaling in glutamatergic inputs to the NAc core contributes to some of the electrophysiological and behavioral hallmarks associated with repeated cocaine administration.

Method for Identifying Small Molecule Inhibitors of the Protein-protein Interaction Between HCN1 and TRIP8b.

Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are expressed ubiquitously throughout the brain, where they function to regulate the excitability of neurons. The subcellular distribution of these channels in pyramidal neurons of hippocampal area CA1 is regulated by tetratricopeptide repeat-containing Rab8b interacting protein (TRIP8b), an auxiliary subunit. Genetic knockout of HCN pore forming subunits or TRIP8b, both lead to an increase in antidepressant-like behavior, suggesting that limiting the function of HCN channels may be useful as a treatment for Major Depressive Disorder (MDD). Despite significant therapeutic interest, HCN channels are also expressed in the heart, where they regulate rhythmicity. To circumvent off-target issues associated with blocking cardiac HCN channels, our lab has recently proposed targeting the protein-protein interaction between HCN and TRIP8b in order to specifically disrupt HCN channel function in the brain. TRIP8b binds to HCN pore forming subunits at two distinct interaction sites, although here the focus is on the interaction between the tetratricopeptide repeat (TPR) domains of TRIP8b and the C terminal tail of HCN1. In this protocol, an expanded description of a method for purifying TRIP8b and executing a high throughput screen to identify small molecule inhibitors of the interaction between HCN and TRIP8b, is described. The method for high throughput screening utilizes a Fluorescence Polarization (FP) -based assay to monitor the binding of a large TRIP8b fragment to a fluorophore-tagged eleven amino acid peptide corresponding to the HCN1 C terminal tail. This method allows 'hit' compounds to be identified based on the change in the polarization of emitted light. Validation assays are then performed to ensure that 'hit' compounds are not artifactual.

Cortical HCN channels: function, trafficking and plasticity.

The hyperpolarization-activated cyclic nucleotide-gated (HCN) channels belong to the superfamily of voltage-gated potassium ion channels. They are, however, activated by hyperpolarizing potentials and are permeable to cations. Four HCN subunits have been cloned, of which HCN1 and HCN2 subunits are predominantly expressed in the cortex. These subunits are principally located in pyramidal cell dendrites, although they are also found at lower concentrations in the somata of pyramidal neurons as well as other neuron subtypes. HCN channels are actively trafficked to dendrites by binding to the chaperone protein TRIP8b. Somato-dendritic HCN channels in pyramidal neurons modulate spike firing and synaptic potential integration by influencing the membrane resistance and resting membrane potential. Intriguingly, HCN channels are present in certain cortical axons and synaptic terminals too. Here, they regulate synaptic transmission but the underlying mechanisms appear to vary considerably amongst different synaptic terminals. In conclusion, HCN channels are expressed in multiple neuronal subcellular compartments in the cortex, where they have a diverse and complex effect on neuronal excitability.

Assessment of the effect of etomidate on voltage-gated sodium channels and action potentials in rat primary sensory cortex pyramidal neurons

Although it is known that general anesthetics can suppress cortical neurons׳ activity, the underlying mechanisms are still poorly understood, especially the kinetic changes of voltage-gated Na+ channels, which are mostly related to neuronal excitability. Some general anesthetics have been reported to affect the voltage-gated Na+ channels in cell culture derived from humans and animals. However no one has ever investigated the effects of etomidate on voltage-gated Na+ channels in pyramidal neurons using a brain slice. The present study uses a whole cell patch-clamp technique to investigate the changes of voltage-gated Na+ channels on primary somatosensory cortex pyramidal neurons under the influence of etomidate. We found that etomidate dose-dependently inhibited Na+ currents of primary somatosensory cortex pyramidal neurons, while shifted the steady-state inactivation curve towards the left and prolonged the recovery time from inactivation. Conversely, etomidate has no effects on the steady-state activation curve. We demonstrated the detailed suppression process of neural voltage-gated Na+ channels by etomidate on slice condition. This may offer new insights into the mechanical explanation for the etomidate anesthesia. Finding the effects of anesthetics on primary somatosensory cortex also provides evidence to help elucidate the potential mechanism by which tactile information integrates during general anesthesia.

Repeated cocaine treatment enhances HIV-1 Tat-induced cortical excitability via over-activation of L-type calcium channels.

The prefrontal cortex (PFC) is dysregulated in neuroAIDS and during cocaine abuse. Repeated cocaine treatment upregulates voltage gated L-type Ca(2+) channels in pyramidal neurons within the rat medial PFC (mPFC). L-type Ca(2+) channels are also upregulated by the HIV-1 neurotoxic protein, Tat, but the role of Tat in pyramidal cell function is unknown. This represents a major knowledge gap as PFC pyramidal neurons are important mediators of behaviors that are disrupted in neuroAIDS and by chronic cocaine exposure. To determine if L-channel-mediated Ca(2+) dysregulation in mPFC pyramidal neurons are a common neuropathogenic site for Tat and chronic cocaine, we evaluated the electrophysiological effects of recombinant Tat on these neurons in forebrain slices taken from rats 1-3days after five, once-daily treatments of cocaine (15mg/kg, ip) or saline. In saline-treated rats, bath-applied Tat facilitated membrane depolarization and firing. Ca(2+) influx was increased (indicated by prolonged Ca(2+) spikes) with low concentrations of Tat (10-40nM), but reduced by higher concentrations (80-160nM), the latter likely reflecting dysfunction associated with excessive excitation. Tat-mediated effects were detected during NMDA/AMPA receptor blockade, and abolished by blocking activated L-channels with diltiazem. In neurons from cocaine-treated rats, the Tat-induced effects on evoked firing and Ca(2+) spikes were significantly enhanced above that obtained with Tat in slices from saline-treated rats. Thus, glutamatergic receptor-independent over-activation of L-channels contributed to the Tat-induced hyper-reactivity of mPFC pyramidal neurons to excitatory stimuli, which was exacerbated in rats repeatedly exposed to cocaine. Such effects may contribute to the exaggerated neuropathology reported for HIV(+) cocaine-abusing individuals.

Enduring cortical alterations after a single in-vivo treatment of HIV-1 Tat.

HIV-1 proteins, including the transactivator of transcription (Tat), are believed to be involved in HIV-associated neurocognitive disorders by disrupting Ca²⁺ homeostasis, which leads to progressive dysregulation, damage, or death of neurons in the brain. We have found previously that bath-applied Tat abnormally increased Ca²⁺ influx through overactivated, voltage-sensitive L-type Ca²⁺ channels in pyramidal neurons within the rat medial prefrontal cortex (mPFC). However, it is unknown whether the Tat-induced Ca²⁺ dysregulation was mediated by increased activity and/or the number of the L-channels. This study tested the hypothesis that transient/early exposure to Tat in vivo promoted enduring L-channel dysregulation in the mPFC without neuron loss. Accordingly, rats were administered a single intracerebroventricular injection of recombinant Tat (80 μg/20 μl; diluted by cerebrospinal fluids to pathophysiological concentrations) or vehicle. Rats were killed 14 days after injection for immunohistochemical assessments of the mPFC, motor cortex, caudate-putamen, and nucleus accumbens. Stereological estimates for positively stained cells indicated a significant increase in the number of cells expressing the pore-forming Ca(v)1.2-α1c subunit of L-channels in the mPFC compared with other regions in Tat-treated or vehicle-treated rat brains. Optical density measurements showed a Tat-induced increase in glial fibrillary acidic protein expression, indicating astrogliosis in the cortical regions. There was no significant loss of neurons in any brain region investigated. These findings indicate that transient Tat exposure in vivo induced enduring L-channel dysregulation and astrogliosis in the mPFC without neuron loss. Such maladaptations may contribute toward dysregulated Ca²⁺ homeostasis and neuropathology in the PFC in the early stages of HIV infection.

Plasma membrane insertion of TRPC5 channels contributes to the cholinergic plateau potential in hippocampal CA1 pyramidal neurons

In cultured hippocampal neurons, transient receptor potential 5 (TRPC5) channels are translocated and inserted into plasma membranes of hippocampal neurons to generate nonselective cation (NSC) currents. We investigated whether TRPC5 channel translocation also contributes to the generation of NSC currents underlying the afterdepolarizations and plateau potentials (PPs) in hippocampal pyramidal cells that are induced by muscarinic receptor activation. Using a biotinylation assay to quantify the change in surface membrane proteins in acute hippocampal slices, we found that muscarinic stimulation significantly enhanced the levels of TRPC5 protein on the membrane surface but not those of TRPC1 or TRPC4 channels. We then investigated the pharmacological sensitivity of the cation current observed during muscarinic stimulation to determine if a component could be due to TRPC5 channels. The TRPC channel antagonists 2-APB and SKF96365 strongly depressed the generation of PPs, the underlying tail currents (I(tail)) and the associated dendritic Ca(2+) influx induced by muscarinic receptor activation in pyramidal neurons. High intracellular concentrations of ATP, which specifically inhibit TRPC5 channels, depressed I(tail). In addition, pretreatment with the calmodulin (CaM) inhibitor W-7, which depresses recombinant TRPC5 currents, inhibited both the cation current (I(tail)) and the surface insertion of TRPC5 channels. Finally, the phosphatidylinositide 3-kinase (PI(3)K) inhibitor wortmannin, which blocks translocation of TRPC5 channels in cell culture, also inhibited both the I(tail) and the surface insertion of TRPC5 channels. Therefore, we conclude that insertion of TRPC5 channels contributes to the generation of the prolonged afterdepolarizations following muscarinic stimulation. This altered plasma membrane expression of TRPC5 channels in pyramidal neurons may play an important role in the generation of prolonged neuronal depolarization and bursting during the epileptiform seizure discharges of epilepsy.

Computational study of hippocampal-septal theta rhythm changes due to β-amyloid-altered ionic channels.

Electroencephagraphy (EEG) of many dementia patients has been characterized by an increase in low frequency field potential oscillations. One of the characteristics of early stage Alzheimer’s disease (AD) is an increase in theta band power (4–7 Hz). However, the mechanism(s) underlying the changes in theta oscillations are still unclear. To address this issue, we investigate the theta band power changes associated with β-Amyloid (Aβ) peptide (one of the main markers of AD) using a computational model, and by mediating the toxicity of hippocampal pyramidal neurons. We use an established biophysical hippocampal CA1-medial septum network model to evaluate four ionic channels in pyramidal neurons, which were demonstrated to be affected by Aβ. They are the L-type Ca2+ channel, delayed rectifying K+ channel, A-type fast-inactivating K+ channel and large-conductance Ca2+-activated K+ channel. Our simulation results demonstrate that only the Aβ inhibited A-type fast-inactivating K+ channel can induce an increase in hippocampo-septal theta band power, while the other channels do not affect theta rhythm. We further deduce that this increased theta band power is due to enhanced synchrony of the pyramidal neurons. Our research may elucidate potential biomarkers and therapeutics for AD. Further investigation will be helpful for better understanding of AD-induced theta rhythm abnormalities and associated cognitive deficits.

Whole cell recording from an organotypic slice preparation of neocortex.

We have been studying the expression and functional roles of voltage-gated potassium channels in pyramidal neurons from rat neocortex. Because of the lack of specific pharmacological agents for these channels, we have taken a genetic approach to manipulating channel expression. We use an organotypic culture preparation (16) in order to maintain cell morphology and the laminar pattern of cortex. We typically isolate acute neocortical slices at postnatal days 8-10 and maintain the slices in culture for 3-7 days. This allows us to study neurons at a similar age to those in our work with acute slices and minimizes the development of exuberant excitatory connections in the slice. We record from visually-identified pyramidal neurons in layers II/III or V using infrared illumination (IR-) and differential interference contrast microscopy (DIC) with whole cell patch clamp in current- or voltage-clamp. We use biolistic (Gene gun) transfection of wild type or mutant potassium channel DNA to manipulate expression of the channels to study their function. The transfected cells are easily identified by epifluorescence microscopy after co-transfection with cDNA for green fluorescent protein (GFP). We compare recordings of transfected cells to adjacent, untransfected neurons in the same layer from the same slice.

Plasticity of L‐type Ca2+ channels after cocaine withdrawal

Increased reactivity of certain frontal cortical brain regions to cocaine re-exposure or drug-associated cues in cocaine-abstinent human addicts is linked to drug craving. Similarly, in rats tested after withdrawal from repeated cocaine exposure, cocaine or other strong excitatory stimuli produce greater activation of pyramidal neurons in the medial prefrontal cortex (mPFC). Our recent findings indicate that the increased mPFC neuronal activation depends primarily upon enhanced voltage-sensitive Ca(2+) influx, most likely through high-voltage activated (HVA) L-type Ca(2+) channels, but the mechanism underlying the enhanced Ca(2+) currents is unknown. In this study, we used a protein crosslinking assay to show that repeated cocaine injections, resulting in behavioral sensitization, increased total protein levels and cell surface expression of HVA-Ca(v)1.2 L-type channels in pyramidal neurons in deep layers of the mPFC. These changes in Ca(v)1.2 L-channels were time dependent and subtype specific (i.e., differed from those observed for Ca(v)1.3 L-channels). Furthermore, we found enhanced PKA activity in the mPFC of cocaine-sensitized rats that persisted for 21 days after withdrawal. PKA phosphorylation of L-channels increases their activity, so Ca(2+) currents after cocaine withdrawal could be enhanced as a result of both increased activity and number of HVA-Ca(v)1.2 L-channels on the cell surface. By increasing the suprafiring threshold excitability of mPFC pyramidal neurons, excessive upregulation of HVA L-channel activity and number may contribute to the cortical hyper-responsiveness that enhances vulnerability to cocaine craving and relapse. More generally, our results are the first to demonstrate that repeated cocaine exposure alters the membrane trafficking of a voltage-sensitive ion channel.

Characterization of Voltage-Gated Ca2+ Conductances in Layer 5 Neocortical Pyramidal Neurons from Rats

Neuronal voltage-gated Ca2+ channels are involved in electrical signalling and in converting these signals into cytoplasmic calcium changes. One important function of voltage-gated Ca2+ channels is generating regenerative dendritic Ca2+ spikes. However, the Ca2+ dependent mechanisms used to create these spikes are only partially understood. To start investigating this mechanism, we set out to kinetically and pharmacologically identify the sub-types of somatic voltage-gated Ca2+ channels in pyramidal neurons from layer 5 of rat somatosensory cortex, using the nucleated configuration of the patch-clamp technique. The activation kinetics of the total Ba2+ current revealed conductance activation only at medium and high voltages suggesting that T-type calcium channels were not present in the patches. Steady-state inactivation protocols in combination with pharmacology revealed the expression of R-type channels. Furthermore, pharmacological experiments identified 5 voltage-gated Ca2+ channel sub-types – L-, N-, R- and P/Q-type. Finally, the activation of the Ca2+ conductances was examined using physiologically derived voltage-clamp protocols including a calcium spike protocol and a mock back-propagating action potential (mBPAP) protocol. These experiments enable us to suggest the possible contribution of the five Ca2+ channel sub-types to Ca2+ current flow during activation under physiological conditions.

Activation of 5-HT2A/C Receptors Counteracts 5-HT1A Regulation of N-Methyl-D-aspartate Receptor Channels in Pyramidal Neurons of Prefrontal Cortex

Abnormal serotonin-glutamate interaction in prefrontal cortex (PFC) is implicated in the pathophysiology of many mental disorders, including schizophrenia and depression. However, the mechanisms by which this interaction occurs remain unclear. Our previous study has shown that activation of 5-HT1A receptors inhibits N-methyl-d-aspartate (NMDA) receptor (NMDAR) currents in PFC pyramidal neurons by disrupting microtubule-based transport of NMDARs. Here we found that activation of 5-HT2A/C receptors significantly attenuated the effect of 5-HT1A on NMDAR currents and microtubule depolymerization. The counteractive effect of 5-HT2A/C on 5-HT1A regulation of synaptic NMDAR response was also observed in PFC pyramidal neurons from intact animals treated with various 5-HT-related drugs. Moreover, 5-HT2A/C stimulation triggered the activation of extracellular signal-regulated kinase (ERK) in dendritic processes. Inhibition of the β-arrestin/Src/dynamin signaling blocked 5-HT2A/C activation of ERK and the counteractive effect of 5-HT2A/C on 5-HT1A regulation of NMDAR currents. Immunocytochemical studies showed that 5-HT2A/C treatment blocked the inhibitory effect of 5-HT1A on surface NR2B clusters on dendrites, which was prevented by cellular knockdown of β-arrestins. Taken together, our study suggests that serotonin, via 5-HT1A and 5-HT2A/C receptor activation, regulates NMDAR functions in PFC neurons in a counteractive manner. 5-HT2A/C, by activating ERK via the β-arrestin-dependent pathway, opposes the 5-HT1A disruption of microtubule stability and NMDAR transport. These findings provide a framework for understanding the complex interactions between serotonin and NMDARs in PFC, which could be important for cognitive and emotional control in which both systems are highly involved.

Output-mode transitions are controlled by prolonged inactivation of sodium channels in pyramidal neurons of subiculum.

Transitions between different behavioral states, such as sleep or wakefulness, quiescence or attentiveness, occur in part through transitions from action potential bursting to single spiking. Cortical activity, for example, is determined in large part by the spike output mode from the thalamus, which is controlled by the gating of low-voltage–activated calcium channels. In the subiculum—the major output of the hippocampus—transitions occur from bursting in the delta-frequency band to single spiking in the theta-frequency band. We show here that these transitions are influenced strongly by the inactivation kinetics of voltage-gated sodium channels. Prolonged inactivation of sodium channels is responsible for an activity-dependent switch from bursting to single spiking, constituting a novel mechanism through which network dynamics are controlled by ion channel gating.

Properties of large conductance calcium-activated potassium channels in pyramidal neurons from the hippocampal CA1 region of adult rats.

The properties of large-conductance Ca(2+)-activated K(+) (BK(Ca)) channels were studied in rat hippocampal CA1 pyramidal neurons by using the patch-clamp technique in the excised-inside-out-patch configuration. The lowest [Ca(2+)](i) in which BK(Ca) channel activities were observed was 0.01 microM with the membrane potential of +20 mV and the [Ca(2+)](i) at which P(O) of the channel is equal to 0.5 was 2 microM. The unitary conductance of the single BK(Ca) channel was 245.4 pS with symmetrical 140 mM K(+) on both sides of the excised membrane. With a fixed [Ca(2+)](i) of 2 microM, P(O) increased e-fold with a 17.0 mV positive change in the membrane potential. Two exponentials, with time constants of 2.8 ms and 19.2 ms at the membrane potential of +120 mV with 2 microM [Ca(2+)](i), were required to describe the observed open time distribution of BK(Ca) channel, suggesting the existence of two distinct open channel states with apparently normal conductance. A BK(Ca) channel occasionally entered an apparent third open channel state with the single channel current amplitude about 45% of the normal amplitude. The properties of BK(Ca) channel, which were found in this study to be more steeply dependent on voltage and more sensitive to [Ca(2+)](i) in adult hippocampal neurons than in cultured or immature hippocampal neurons, may be responsible for the shortened duration of action potential in hippocampal CA1 pyramidal neurons of adult rat.

Long-term potentiation of Ca2+ signal in the rat auditory cortex.

The Ca2+ signal in supragranular layers of the rat auditory cortex (AC) was studied in slice preparations using rhod-2, a Ca2+ indicator. White matter stimulation elicited an increase in the Ca2+ signal, which was maximal in the image taken 34 ms after stimulation. This peak time was the same as that of the Ca2+ signal in pyramidal neurons injected with rhod-2. The intensity of the Ca2+ signal was proportional to the amplitude of the field potentials in supragranular layers. The Ca2+ signal was inhibited almost completely by 200 microM Ni2+ , but only slightly by 50 microM D-2-amino-5-phosphonovalerate (APV), an NMDA-receptor antagonist. Tetanic stimulation of the white matter or supragranular layers elicited long-term potentiation (LTP) of the Ca2+ signal in AC slices, but the potentiation was not clear in slices of the visual cortex (VC). The induction of LTP of the field potentials in AC slices was blocked by 50 microM APV or 50 microM Ni2+. These results indicate that Ca2+ influx through Ni2+ -sensitive Ca2+ channels in pyramidal neurons is potentiated by tetanic stimulation in parallel with LTP of neural activities and might be important for the induction of LTP in AC slices.