Dataset Information

Fluctuation Analysis in Nonstationary Conditions: Single Ca2+ Channel Current in Pyramidal Neurons.

ABSTRACT: Fluctuation analysis is a method that allows measurement of the single-channel current of ion channels even when it is too small to be resolved directly with the patch-clamp technique. This is the case for voltage-gated calcium channels. They are present in all mammalian central neurons, controlling presynaptic release of transmitter, postsynaptic signaling, and synaptic integration. The amplitudes of their single-channel currents in a physiological concentration of extracellular calcium, however, are small and not well determined. But measurement of this quantity is essential for estimating numbers of functional voltage-gated calcium channels in the membrane and the size of channel-associated calcium signaling domains, and for understanding the stochastic nature of calcium signaling. Here, we recorded the voltage-gated calcium channel current in nucleated patches from layer 5 pyramidal neurons in rat neocortex, in physiological external calcium (1-2 mM). The ensemble-averaging of current responses required for conventional fluctuation analysis proved impractical because of the rapid rundown of calcium channel currents. We therefore developed a more robust method, using mean current fitting of individual current responses and band-pass filtering. Furthermore, voltage-ramp stimulation proved useful. We validated the accuracy of the method by analyzing simulated data. At an external calcium concentration of 1 mM, and a membrane potential of -20 mV, we found that the average single-channel current amplitude was ?0.04 pA, increasing to 0.065 pA at 2 mM external calcium, and 0.12 pA at 5 mM. The relaxation time constant of the fluctuations was in the range 0.2-0.8 ms. The results are relevant to understanding the stochastic properties of dendritic Ca2+ spikes in neocortical layer 5 pyramidal neurons. With the reported method, single-channel current amplitude of native voltage-gated calcium channels can be resolved accurately despite conditions of unstable rundown.

SUBMITTER: Scheppach C

PROVIDER: S-EPMC5738524 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Publications

Fluctuation Analysis in Nonstationary Conditions: Single Ca<sup>2+</sup> Channel Current in Pyramidal Neurons.

Scheppach Christian C Robinson Hugh P C HPC

Biophysical journal 20171201 11

Fluctuation analysis is a method that allows measurement of the single-channel current of ion channels even when it is too small to be resolved directly with the patch-clamp technique. This is the case for voltage-gated calcium channels. They are present in all mammalian central neurons, controlling presynaptic release of transmitter, postsynaptic signaling, and synaptic integration. The amplitudes of their single-channel currents in a physiological concentration of extracellular calcium, howeve ...[more]

PMID: 29211992

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Project description:Oxytocin (OXT) and OXT receptor (OXTR)-mediated signaling control excitability, firing patterns, and plasticity of hippocampal CA2 pyramidal neurons, which are pivotal in generation of brain oscillations and social memory. Nonetheless, the ionic mechanisms underlying OXTR-induced effects in CA2 neurons are not fully understood. Using slice physiology in a reporter mouse line and interleaved current-clamp and voltage-clamp experiments, we systematically identified the ion channels modulated by OXT signaling in CA2 pyramidal cells (PYRs) in mice of both sexes and explored how changes in channel conductance support altered electrical activity. Activation of OXTRs inhibits an outward potassium current mediated by inward rectifier potassium channels (I Kir) and thus favoring membrane depolarization. Concomitantly, OXT signaling also diminishes inward current mediated by hyperpolarization-activated cyclic-nucleotide-gated (HCN) channels (I h), providing a hyperpolarizing drive. The combined reduction in both I Kir and I h synergistically elevate the membrane resistance and favor dendritic integration while the membrane potential is restrained from quickly depolarizing from rest. As a result, the responsiveness of CA2 PYRs to synaptic inputs is highly sharpened during OXTR activation. Unexpectedly, OXTR signaling also strongly enhances a tetrodotoxin-resistant (TTX-R), voltage-gated sodium current that helps drive the membrane potential to spike threshold and thus promote rhythmic firing. This novel array of OXTR-stimulated ionic mechanisms operates in close coordination and underpins OXT-induced burst firing, a key step in CA2 PYRs' contribution to hippocampal information processing and broader influence on brain circuitry. Our study deepens our understanding of underpinnings of OXT-promoted social memory and general neuropeptidergic control of cognitive states.SIGNIFICANCE STATEMENT Oxytocin (OXT) plays key roles in reproduction, parenting and social and emotional behavior, and deficiency in OXT receptor (OXTR) signaling may contribute to neuropsychiatric disorders. We identified a novel array of OXTR-modulated ion channels that operate in close coordination to retune hippocampal CA2 pyramidal neurons, enhancing responsiveness to synaptic inputs and sculpting output. OXTR signaling inhibits both potassium conductance (I Kir) and mixed cation conductance (I h), engaging opposing influences on membrane potential, stabilizing it while synergistically elevating membrane resistance and electrotonic spread. OXT signaling also facilitates a tetrodotoxin-resistant (TTX-R) Na+ current, not previously described in hippocampus (HP), engaged on further depolarization. This TTX-R current lowers the spike threshold and supports rhythmic depolarization and burst firing, a potent driver of downstream circuitry.

Dataset Information

Fluctuation Analysis in Nonstationary Conditions: Single Ca2+ Channel Current in Pyramidal Neurons.

Publications

Fluctuation Analysis in Nonstationary Conditions: Single Ca<sup>2+</sup> Channel Current in Pyramidal Neurons.

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