A persistent sodium current in acutely isolated histaminergic neurons from rat hypothalamus

doi:10.1016/0306-4522(94)00593-T

Neuroscience

Volume 66, Issue 1, May 1995, Pages 143-149

https://doi.org/10.1016/0306-4522(94)00593-T Get rights and content

Abstract

Histamine neurons acutely dissociated from the tuberomammillary nucleus of the rat hypothalamus were studied in whole-cell and cell-attached patch-clamp experiments. Electrophysiological properties of dissociated cells were found to be similar to those recorded in slice experiments using microelectrodes. Tuberomammillary neurons fired spontaneously and this activity persisted when Cs⁺ (1.5 mM) was added to, or when K⁺ was removed from the extracellular solution. In whole-cell experiments a persistent tetrodotoxin-sensitive inward current was recorded. In cell attached recordings voltage-gated sodium channels displayed either normal or non-inactivating behavior.

These results provide a further analysis of the properties of histaminergic neurons and indicate that spontaneous activity is intrinsic to individual neurons. Evidence for a non-inactivating tetrodotoxin-sensitive sodium current is presented. Single channel recordings indicate that this current is the result of non-inactivating behavior of sodium channels. Such a current is well suited for biasing tuberomammillary neurons toward spontaneous activity.

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Sequence variations at I260 and A1731 contribute to persistent currents in Drosophila sodium channels
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Citation Excerpt :
Besides the transient (i.e., fast inactivating) sodium currents (INaT), which are responsible for the upstroke of action potentials, there are also tetrodotoxin (TTX)-sensitive non-inactivating or persistent currents (INaP), which activate at subthreshold voltages and cannot inactivate completely even with prolonged depolarization (Taylor, 1993; Crill, 1996; Stafstrom, 2007; Kiss, 2008). INaP have been detected in numerous types of vertebrate neurons in the brain, such as the suprachiasmatic nucleus (Pennartz et al., 1997; Jackson et al., 2004); cerebellar nuclei (Raman et al., 2000); and tuberomammillary neurons (Llinas and Alonso, 1992; Uteshev et al., 1995; Taddese and Bean, 2002). INaP are also found in invertebrate neurons, such as squid giant axons (Rakowski et al., 2002; Clay, 2003), squid olfactory receptor neurons (Chen and Lucero, 1999), leech spontaneously active heart interneurons (Opdyke and Calabrese, 1994), Drosophila aCC/RP2 motor neurons (Mee et al., 2004), and cockroach terminal abdominal efferent dorsal unpaired median (DUM) octopaminergic neurons (Lapied et al., 1990).
Tetrodotoxin-sensitive persistent sodium currents, I_NaP, that activate at subthreshold voltages, have been detected in numerous vertebrate and invertebrate neurons. These currents are believed to be critical for regulating neuronal excitability. However, the molecular mechanism underlying I_NaP is controversial. In this study, we identified an I_NaP with a broad range of voltage dependence, from −60 mV to 20 mV, in a Drosophila sodium channel variant expressed in Xenopus oocytes. Mutational analysis revealed that two variant-specific amino acid changes, I260T in the S4–S5 linker of domain I (ILS4–S5) and A1731V in the voltage sensor S4 of domain IV (IVS4), contribute to the I_NaP. I260T is critical for the portion of I_NaP at hyperpolarized potentials. The T260-mediated I_NaP is likely the result of window currents flowing in the voltage range where the activation and inactivation curves overlap. A1731V is responsible for impaired inactivation and contributes to the portion of I_NaP at depolarized potentials. Furthermore, A1731V causes enhanced activity of two site-3 toxins which induce persistent currents by inhibiting the outward movement of IVS4, suggesting that A1731V inhibits the outward movement of IVS4. These results provided molecular evidence for the involvement of distinct mechanisms in the generation of I_NaP: T260 contributes to I_NaP via enhancement of the window current, whereas V1731 impairs fast inactivation probably by inhibiting the outward movement of IVS4.
Evidence for the exclusive expression of functional homomeric α7 nAChRs in hypothalamic histaminergic tuberomammillary neurons in rats
2014, Neuroscience Letters
Citation Excerpt :
Its ventral group is located on the edge of the posterior hypothalamus forming several large clusters of cells that can be easily identified and dissociated using a phase-contrast microscope (Fig. 1) [31,32]. These clusters act as a convenient source of acutely dissociated cells characterized by uniform neuronal and nAChR properties similar to those observed in in vivo and ex vivo preparations [10,25,27–29,31–35]. Therefore, TM neurons have been previously proposed as a useful source of native α7-containing nAChRs.
Hypothalamic histaminergic tuberomammillary (TM) neurons in rats express high densities of nicotinic acetylcholine receptors (nAChRs) whose Ca²⁺ permeability, kinetic and pharmacological properties are similar to those of heterologous homomeric α7 nAChRs. However, native α7 nAChR subunits can co-assemble with β or α5 nAChR subunits to form functional heteromeric α7-containing α7β or α7α5 nAChRs with kinetics and pharmacology similar to those of α7 homomers. Therefore, although TM nAChRs have been used as an ex vivo model of functional α7 homomers, the molecular makeup of TM nAChRs has not been determined and the expression of functional α7-containing heteromers in TM neurons has not been excluded. To determine the profile of TM nAChR subunit transcripts, we have conducted single-cell qRT-PCR experiments using acutely dissociated TM neurons in rats. TM neurons were found to express transcripts of only principal α3, α6 and α7 nAChR subunits. Transcripts of other known mammalian neuronal subunits (α2, α4–5, α9–10, β2–4) were not detected. In the absence of β and α5 subunits, the expression of functional α7-containing heteromers in TM neurons is highly unlikely because principal α3, α6 and α7 nAChR subunits alone are not known to form functional heteromeric nAChRs. These results support the exclusive expression of native functional α7 homomers in rat TM neurons and introduce these neurons as a unique reliable source of native functional homomeric α7 nAChRs suitable for ex vivo and in vitro pharmacological assays in developing selective α7 nAChR agents.
Network Experimental Approaches: Ex vivo Recording
2014, Neuronal Networks in Brain Function, CNS Disorders, and Therapeutics
The cellular and molecular mechanisms supporting cognitive and autonomic homeostases can be effectively investigated in ex vivo preparations (e.g. acute brain sections) where they are disconnected from the behavioral context and where parameters that define physiological processes can be manipulated and optimized. Therefore, ex vivo preparations allow modeling of pathophysiological conditions and delineation of the properties and performance of functionally defined neuronal circuits. However, replacing a functional brain with brain sections may limit the interpretation and physiological relevance of results because surgical manipulations damage brain tissues, while the artificial cerebrospinal fluid only approximately emulates the native physiological milieu (see Chapter 10). This chapter reasons that a compromise between the benefits and limitations of ex vivo preparations can be achieved to provide insights into network mechanisms that are not adequately addressed by in vivo (e.g. freely moving or anesthetized animals) or in vitro (e.g. cultured cells and cell lines) experimental approaches. Three distinct mechanisms (i.e. direct synchronization, inhibition of asynchrony, and enhancement of neuronal excitability) that enhance the impact of individual neurons on the network function are discussed using experimental data obtained from the olfactory, hippocampal, and hypothalamic acute brain preparations.
Cholinergic modulation of neurons in the gustatory region of the nucleus of the solitary tract
2006, Brain Research
Citation Excerpt :
Ih has been shown to act as a pacemaker current in Purkinje fibers of the heart (DiFrancesco, 1981) and in thalamic neurons (McCormick and Pape, 1990). However, in histaminergic tuberomammillary neurons, the influence of Ih on neuronal spontaneous firing is minimal (Kamondi and Reiner, 1991; Uteshev et al., 1995). Activation of Ih at membrane potentials near the rNTS resting potential (i.e., −65 mV) would be expected to be limited, although its influence on rNTS spontaneous firing cannot be ruled out because various neurotransmitters (dopamine, noradrenalin, and serotonin) have been shown to alter the activation curve of Ih, making its effects on spontaneous firing possible even at the resting membrane potential (Bobker and Williams, 1989; Pape and McCormick, 1989; Vargas and Lucero, 1999; Wu and Hablitz, 2005).
The rostral portion of the nucleus of the solitary tract (rNST) is an obligatory relay for gustatory afferent input on its way to the forebrain. Previous studies have demonstrated excitation of rNTS neurons by glutamate and substance P and inhibition by γ-aminobutyric acid (GABA) and met-enkephalin (ENK). Despite the existence of cholinergic neurons and putative terminals within the rNTS, there are no data on the effects of acetylcholine (ACh) on rNTS processing. Here, we use patch-clamp recording of rNTS neurons in vitro to examine ACh-mediated responses and voltage-gated conductances in these cells. Results revealed (1) intrinsic voltage-gated inhibition via activation of voltage-gated potassium A-channels (I_A), found almost exclusively in the medial rNTS, and hyperpolarization-activated potassium/sodium channels (I_h), found more frequently in the lateral rNST; and (2) ligand-gated inhibition via activation of muscarinic m2 ACh receptors (mAChRs) linked to inward rectifier potassium channels (K_ir) evenly distributed throughout the rNTS, a mechanism dependent on cholinergic inputs. Muscarinic responses were blocked by AFDX-116, a selective m2 mAChR antagonist, and by BaCl₂, an antagonist of K_ir channels. In addition, many rNTS neurons exhibited excitation via α7 and non-α7 nicotinic AChRs. Non-α7 nAChRs, blocked by 10 μM mecamylamine, occurred more frequently in the lateral rNTS. In contrast, α7 nAChRs, blocked by 20 nM methyllcaconitine, were evenly distributed across the nucleus. As previously reported for voltage-activated conductances, none of these currents was related to neuronal morphology. These voltage- and ligand-dependent inhibitory mechanisms would be expected to contribute to the modulation of gustatory processing through the NST.
Decreases in CaMKII activity trigger persistent potentiation of intrinsic excitability in spontaneously firing vestibular nucleus neurons
2005, Neuron
Calcium/calmodulin-dependent protein kinase II (CaMKII) has been described as a biochemical switch that is turned on by increases in intracellular calcium to mediate synaptic plasticity. Here, we show that reductions in CaMKII activity trigger persistent increases in intrinsic excitability. In spontaneously firing vestibular nucleus neurons, CaMKII activity is near maximal, and blockade of CaMKII activity increases excitability by reducing BK-type calcium-activated potassium currents. Firing rate potentiation, a form of plasticity in which synaptic inhibition induces long-lasting increases in excitability, is occluded by prior blockade of CaMKII and blocked by addition of constitutively active CaMKII. Reductions in CaMKII activity are necessary and sufficient to induce firing rate potentiation and may contribute to motor learning in the vestibulo-ocular reflex.
Somatic Ca<sup>2+</sup> dynamics in response to choline-mediated excitation in histaminergic tuberomammillary neurons
2005, Neuroscience
Histaminergic tuberomammillary (TM) neurons of the posterior hypothalamus have been implicated in cognition, alertness and sleep-wakefulness cycles. Spontaneous firing of TM neurons has been associated with histamine release and wakefulness. The expression of α7 nicotinic acetylcholine receptors (nAChRs) in TM neurons suggests a role for endogenous choline and for nicotinic drugs in the regulation of intracellular Ca²⁺ metabolism, normal TM neuronal activity and histamine release. First, we established the link between TM neuronal spontaneous firing frequency and cytosolic free Ca²⁺ concentration ([Ca²⁺]_i). A strong correlation was observed: an onset of spontaneous firing (3–4Hz) was accompanied by a 20-fold increase in [Ca²⁺]_i from 56±18nM to 1.0±0.6μM. The same range of firing frequencies has been observed in TM neurons in vivo and is associated with wakefulness. Secondly, choline-induced activation of α7 nAChRs did not elevate [Ca²⁺]_i directly, i.e. in the absence of high-threshold voltage-gated Ca²⁺ channel (HVGCC) activation. Cd²⁺ (200μM) completely blocked all Ca²⁺ signals, but inhibited only 37±16% of α7 nAChR-mediated currents. Thirdly, the responsiveness of [Ca²⁺]_i to choline-mediated excitation was inhibited by hyperpolarization and enhanced by depolarization, sensitizing [Ca²⁺]_i at membrane voltages associated with normal TM neuronal activity. These properties of [Ca²⁺]_i define the ability of TM neurons to translate cholinergic stimuli of identical strengths into different cytosolic Ca²⁺ effects, providing the physiological substrate for state-specific modulation of incoming cholinergic information and would be expected to play a very important role in determining activity profiles of TM neurons exposed to elevated concentrations of cholinergic agents, such as choline and nicotine.

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^*: Present address: Department of Physiology, University of Toronto, Toronto, Ontario, Canada, M5S 1A8.

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A persistent sodium current in acutely isolated histaminergic neurons from rat hypothalamus

Abstract

Brain Res.

Prog. Neurobiol.

Brain Res.

Neuron

Biophys. J.

Neurosci. Lett.

Brain Res.

Trends Neurosci.

Brain Res.

The histaminergic system in the guinea pig central nervous system: an immunocytochemical mapping study using an antiserum against histamine

J. comp. Neurol.

Modal gating of Na+ channels as a mechanism of persistent Na+ current in pyramidal neurons from rat and cat sensorimotor cortex

J. Neurosci.

Modal gating of Na⁺ channels as a mechanism of persistent Na⁺ current in pyramidal neurons from rat and cat sensorimotor cortex