Carrier-Mediated GABA Release Activates GABA Receptors on Hippocampal Neurons

Gaspary, Heidi L., Wengang Wang, and George B. Richerson. Carrier-mediated GABA release activates GABA receptors on hippocampal neurons. J. Neurophysiol. 80: 270–281, 1998. γ-Aminobutyric acid (GABA) transporters are electrogenic and sodium-dependent and can operate in reverse when cells are depolarized or when there is reversal of the inward sodium gradient. However, the functional relevance of this phenomenon is unclear. We have examined whether depolarization induced by a physiologically relevant increase in extracellular [K+] leads to sufficient amounts of carrier-mediated GABA release to activate GABAA receptors on neurons. Patch-clamp recordings were made from rat hippocampal neurons in culture with solutions designed to isolate chloride currents in the recorded neuron. Pressure microejection was used to increase extracellular [K+] from 3 to 12 mM. After blockade of vesicular GABA release by removal of extracellular calcium, this stimulus induced a large conductance increase in hippocampal neurons [18.9 ± 6.8 (SD) nS; n = 16]. This was blocked by the GABAA receptor antagonists picrotoxin and bicuculline and had a reversal potential that followed the Nernst potential for chloride, indicating that it was mediated by GABAA receptor activation. Similar responses occurred after block of vesicular neurotransmitter release by tetanus toxin. GABAA receptors also were activated when an increase in extracellular [K+] (from 3 to 13 mM) was combined with a reduction in extracellular [Na+] or when cells were exposed to a decrease in extracellular [Na+] alone. These results indicate that depolarization and/or reversal of the Na+ gradient activated GABA receptors via release of GABA from neighboring cells. We found that the GABA transporter antagonists 1-(4,4-diphenyl-3-butenyl)-3-piperidinecarboxylic acid hydrochloride (SKF89976A; 20–100 μM) and 1-(2-{[(diphenylmethylene)amino]oxy}ethyl) -1, 2, 5, 6 - tetrahydro - 3 - pyridine - carboxylic acid hydrochloride (NO-711; 10 μM) both decreased the responses, indicating that the release of GABA resulted from reversal of the GABA transporter. We propose that carrier-mediated GABA release occurs in vivo during high-frequency neuronal firing and seizures, and dynamically modulates inhibitory tone.

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Vigabatrin Induces Tonic Inhibition Via GABA Transporter Reversal Without Increasing Vesicular GABA Release

Journal of Neurophysiology ◽

10.1152/jn.00856.2002 ◽

2003 ◽

Vol 89 (4) ◽

pp. 2021-2034 ◽

Cited By ~ 101

Author(s):

Yuanming Wu ◽

Wengang Wang ◽

George B. Richerson

Keyword(s):

Patch Clamp ◽

Gaba Receptors ◽

Hippocampal Neurons ◽

Gaba Release ◽

Tonic Inhibition ◽

Gabaergic Inhibition ◽

Gaba Transporter ◽

Whole Cell ◽

Ambient Gaba

Two forms of GABAergic inhibition coexist: fast synaptic neurotransmission and tonic activation of GABA receptors due to ambient GABA. The mechanisms regulating ambient GABA have not been well defined. Here we examined the role of the GABA transporter in the increase in ambient [GABA] induced by the anticonvulsant vigabatrin. Pretreatment of cultured rat hippocampal neurons with vigabatrin (100 μM) for 2–5 days led to a large increase in ambient [GABA] that was measured as the change in holding current induced by bicuculline during patch-clamp recordings. In contrast, there was a decrease in the frequency of spontaneous miniature inhibitory postsynaptic currents mIPSCs with no change in their amplitude distribution, and a decrease in the magnitude of IPSCs evoked by presynaptic stimulation during paired recordings. The increase in ambient [GABA] was not prevented by blockade of vesicular GABA release with tetanus toxin or removal of extracellular calcium. During perforated patch recordings, the increase in ambient [GABA] was prevented by blocking the GABA transporter, indicating that the GABA transporter was continuously operating in reverse and releasing GABA. In contrast, blocking the GABA transporter increased ambient [GABA] during whole cell patch-clamp recordings unless GABA and Na+ were added to the recording electrode solution, indicating that whole cell recordings can lead to erroneous conclusions about the role of the GABA transporter in control of ambient GABA. We conclude that the equilibrium for the GABA transporter is a major determinant of ambient [GABA] and tonic GABAergic inhibition. We propose that fast GABAergic neurotransmission and tonic inhibition can be independently modified and play complementary roles in control of neuronal excitability.

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GABA Transporter-1 (GAT1)-Deficient Mice: Differential Tonic Activation of GABAA Versus GABAB Receptors in the Hippocampus

Journal of Neurophysiology ◽

10.1152/jn.00240.2003 ◽

2003 ◽

Vol 90 (4) ◽

pp. 2690-2701 ◽

Cited By ~ 149

Author(s):

Kimmo Jensen ◽

Chi-Sung Chiu ◽

Irina Sokolova ◽

Henry A. Lester ◽

Istvan Mody

Keyword(s):

Gaba Release ◽

Gabab Receptors ◽

Acid Decarboxylase ◽

Wild Type ◽

Gaba Transporter ◽

Inhibitory Neurotransmitter ◽

Gaba Transporters ◽

Gaba Transporter 1 ◽

Glutamic Acid Decarboxylase 65 ◽

Deficient Mice

After its release from interneurons in the CNS, the major inhibitory neurotransmitter GABA is taken up by GABA transporters (GATs). The predominant neuronal GABA transporter GAT1 is localized in GABAergic axons and nerve terminals, where it is thought to influence GABAergic synaptic transmission, but the details of this regulation are unclear. To address this issue, we have generated a strain of GAT1-deficient mice. We observed a large increase in a tonic postsynaptic hippocampal GABAA receptor-mediated conductance. There was little or no change in the waveform or amplitude of spontaneous inhibitory postsynaptic currents (IPSCs) or miniature IPSCs. In contrast, the frequency of quantal GABA release was one-third of wild type (WT), although the densities of GABAA receptors, GABAB receptors, glutamic acid decarboxylase 65 kDa, and vesicular GAT were unaltered. The GAT1-deficient mice lacked a presynaptic GABAB receptor tone, present in WT mice, which reduces the frequency of spontaneous IPSCs. We conclude that GAT1 deficiency leads to enhanced extracellular GABA levels resulting in an overactivation of GABAA receptors responsible for a postsynaptic tonic conductance. Chronically elevated GABA levels also downregulate phasic GABA release and reduce presynaptic signaling via GABAB receptors thus causing an enhanced tonic and a diminished phasic inhibition.

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GAT-1 and Reversible GABA Transport in Bergmann Glia in Slices

Journal of Neurophysiology ◽

10.1152/jn.2002.88.3.1407 ◽

2002 ◽

Vol 88 (3) ◽

pp. 1407-1419 ◽

Cited By ~ 57

Author(s):

L. Barakat ◽

A. Bordey

Keyword(s):

Gaba Receptors ◽

Receptor Activation ◽

Bergmann Glia ◽

Gaba Uptake ◽

Induced Voltage ◽

Nipecotic Acid ◽

Gaba Transporters ◽

Whole Cell Patch Clamp ◽

Inward Currents

Although glial GABA uptake and release have been studied in vitro, GABA transporters (GATs) have not been characterized in glia in slices. Whole cell patch-clamp recordings were obtained from Bergmann glia in rat cerebellar slices to characterize carrier-mediated GABA influx and efflux. GABA induced inward currents at −70 mV that could be pharmacologically separated into GABAA receptor and GAT currents. In the presence of GABAA/B/C receptor blockers, mean GABA-induced currents measured −48 pA at −70 mV, were inwardly rectifying between −70 and +50 mV, were inhibited by external Na+ removal, and were diminished by reduction of external Cl−. Nontransportable blockers of GAT-1 (SKF89976-A and NNC-711) and a transportable blocker of all the GAT subtypes (nipecotic acid) reversibly reduced GABA-induced transport currents by 68 and 100%, respectively. A blocker of BGT-1 (betaine) had no effect. SKF89976-A and NNC-711 also suppressed baseline inward currents that likely result from tonic GAT activation by background GABA. The substrate agonists, nipecotic acid and β-alanine but not betaine, induced voltage- and Na+-dependent currents. With Na+ and GABA inside the patch pipette or intracellular GABA perfusion during the recording, SKF89976-A blocked baseline outward currents that activated at −60 mV and increased with more depolarized potentials. This carrier-mediated GABA efflux induced a local accumulation of extracellular GABA detected by GABAA receptor activation on the recorded cell. Overall, these results indicate that Bergmann glia express GAT-1 that are activated by ambient GABA. In addition, GAT-1 in glia can work in reverse and release sufficient GABA to activate nearby GABA receptors.

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Resonance with subthreshold oscillatory drive organizes activity and optimizes learning in neural networks

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1716933115 ◽

2018 ◽

Vol 115 (13) ◽

pp. E3017-E3025 ◽

Cited By ~ 11

Author(s):

James P. Roach ◽

Aleksandra Pidde ◽

Eitan Katz ◽

Jiaxing Wu ◽

Nicolette Ognjanovski ◽

...

Keyword(s):

Hippocampal Neurons ◽

External Input ◽

Neuronal Firing ◽

Network Activity ◽

Dependent Manner ◽

Information Encoding ◽

Subthreshold Oscillations ◽

Neuronal Network Model ◽

And Performance

Network oscillations across and within brain areas are critical for learning and performance of memory tasks. While a large amount of work has focused on the generation of neural oscillations, their effect on neuronal populations’ spiking activity and information encoding is less known. Here, we use computational modeling to demonstrate that a shift in resonance responses can interact with oscillating input to ensure that networks of neurons properly encode new information represented in external inputs to the weights of recurrent synaptic connections. Using a neuronal network model, we find that due to an input current-dependent shift in their resonance response, individual neurons in a network will arrange their phases of firing to represent varying strengths of their respective inputs. As networks encode information, neurons fire more synchronously, and this effect limits the extent to which further “learning” (in the form of changes in synaptic strength) can occur. We also demonstrate that sequential patterns of neuronal firing can be accurately stored in the network; these sequences are later reproduced without external input (in the context of subthreshold oscillations) in both the forward and reverse directions (as has been observed following learning in vivo). To test whether a similar mechanism could act in vivo, we show that periodic stimulation of hippocampal neurons coordinates network activity and functional connectivity in a frequency-dependent manner. We conclude that resonance with subthreshold oscillations provides a plausible network-level mechanism to accurately encode and retrieve information without overstrengthening connections between neurons.

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Amyloid-β Inhibits PDGFβ Receptor Activation and Prevents PDGF-BBInduced Neuroprotection

Current Alzheimer Research ◽

10.2174/1567205015666180110110321 ◽

2018 ◽

Vol 15 (7) ◽

pp. 618-627 ◽

Cited By ~ 2

Author(s):

Hui Liu ◽

Golam T. Saffi ◽

Maryam S. Vasefi ◽

Youngjik Choi ◽

Jeff S. Kruk ◽

...

Keyword(s):

Intracellular Signaling ◽

Hippocampal Neurons ◽

Neuronal Damage ◽

Neuroblastoma Cell ◽

Amyloid Β ◽

Receptor Activation ◽

Cell Number ◽

Physical Interaction ◽

Protective Effects

Background: PDGFβ receptors and their ligand, PDGF-BB, are upregulated in vivo after neuronal insults such as ischemia. When applied exogenously, PDGF-BB is neuroprotective against excitotoxicity and HIV proteins. Objective: Given this growth factor's neuroprotective ability, we sought to determine if PDGF-BB would be neuroprotective against amyloid-β (1-42), one of the pathological agents associated with Alzheimer's disease (AD). Methods and Results: In both primary hippocampal neurons and the human-derived neuroblastoma cell line, SH-SY5Y, amyloid-β treatment for 24 h decreased surviving cell number in a concentrationdependent manner. Pretreatment with PDGF-BB failed to provide any neuroprotection against amyloid-β in primary neurons and only very limited protective effects in SH-SY5Y cells. In addition to its neuroprotective action, PDGF promotes cell growth and division in several systems, and the application of PDGFBB alone to serum-starved SH-SY5Y cells resulted in an increase in cell number. Amyloid-β attenuated the mitogenic effects of PDGF-BB, inhibited PDGF-BB-induced PDGFβ receptor phosphorylation, and attenuated the ability of PDGF-BB to protect neurons against NMDA-induced excitotoxicity. Despite the ability of amyloid-β to inhibit PDGFβ receptor activation, immunoprecipitation experiments failed to detect a physical interaction between amyloid-β and PDGF-BB or the PDGFβ receptor. However, G protein-coupled receptor transactivation of the PDGFβ receptor (an exclusively intracellular signaling pathway) remained unaffected by the presence of amyloid-β. Conclusions: As the PDGF system is upregulated upon neuronal damage, the ability of amyloid-β to inhibit this endogenous neuroprotective system should be further investigated in the context of AD pathophysiology.

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GABAergic Modulation of the Activity of Globus Pallidus Neurons in Primates: In Vivo Analysis of the Functions of GABA Receptors and GABA Transporters

Journal of Neurophysiology ◽

10.1152/jn.00068.2005 ◽

2005 ◽

Vol 94 (2) ◽

pp. 990-1000 ◽

Cited By ~ 50

Author(s):

Adriana Galvan ◽

Rosa M. Villalba ◽

Sara M. West ◽

Nigel T. Maidment ◽

Larry C. Ackerson ◽

...

Keyword(s):

Globus Pallidus ◽

Gaba Receptors ◽

Synaptic Cleft ◽

Considerable Proportion ◽

Electron Microscopic ◽

Gaba Transporters ◽

Endogenous Gaba ◽

In Vivo Analysis ◽

Gabaergic Modulation

Neurons in the external and internal segment of the globus pallidus (GPe and GPi, respectively) receive substantial GABAergic inputs from the striatum and through axon collaterals of neighboring pallidal neurons. The effects of GABA on pallidal activity depend on the synaptic localization of GABA receptors and the distribution and activity of GABA transporters (GATs). To explore the contribution of GABA receptors and transporters to pallidal function, we recorded the activity of single neurons in GPe or GPi before, during, and after local microinjections of GABAergic compounds in awake rhesus monkeys. Activation of GABAA or GABAB receptors with muscimol or baclofen, respectively, inhibited pallidal activity. These effects were reversed by concomitant infusion of the respective GABA receptor antagonists, gabazine and CGP-55845. Given alone, the antagonists were without consistent effect. Application of the selective GAT-1 inhibitor, SKF-89976A, and the semiselective GAT-3 blocker, SNAP-5114, decreased pallidal activity. Both GAT inhibitors increased GABA levels in the pallidum, as measured by microdialysis. Electron microscopic observations revealed that these transporters are located on glial processes and unmyelinated axonal segments, but rarely on terminals. Our results indicate that activation of GABAA and GABAB receptors inhibits neuronal activity in both segments of the pallidum. GAT-1 and GAT-3 are involved in the modulation of endogenous GABA levels and may be important in regulating the extrasynaptic levels of GABA. Together with previous evidence that a considerable proportion of pallidal GABA receptors are located outside the synaptic cleft, our experiments strongly support the importance of extrasynaptic GABAergic transmission in the primate pallidum.

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Rapid regulation of tonic GABA currents in cultured rat hippocampal neurons

Journal of Neurophysiology ◽

10.1152/jn.00460.2012 ◽

2013 ◽

Vol 109 (3) ◽

pp. 803-812 ◽

Cited By ~ 11

Author(s):

Christopher B. Ransom ◽

Wucheng Tao ◽

Yuanming Wu ◽

William J. Spain ◽

George B. Richerson

Keyword(s):

Hippocampal Neurons ◽

Brain Slices ◽

Reversal Potential ◽

Gaba Release ◽

Tonic Inhibition ◽

Concanamycin A ◽

Voltage Dependent ◽

Tonic Current ◽

Induced Changes ◽

Time Frames

Subacute and chronic changes in tonic GABAergic inhibition occur in human and experimental epilepsy. Less is known about how tonic inhibition is modulated over shorter time frames (seconds). We measured endogenous tonic GABA currents from cultured rat hippocampal neurons to evaluate how they are affected by 1) transient increases in extracellular GABA concentration ([GABA]), 2) transient postsynaptic depolarization, and 3) depolarization of presynaptic cells. Transient increases in [GABA] (1 μM) reduced tonic currents; this reduction resulted from GABA-induced shifts in the reversal potential for GABA currents ( EGABA). Transient depolarization of postsynaptic neurons reversed the effects of exogenous GABA and potentiated tonic currents. The voltage-dependent potentiation of tonic GABA currents was independent of EGABA shifts and represented postdepolarization potentiation (PDP), an intrinsic GABAA receptor property (Ransom CB, Wu Y, Richerson GB. J Neurosci 30: 7672–7684, 2010). Inhibition of vesicular GABA release with concanamycin A (ConA) did not affect tonic currents. In ConA-treated cells, transient application of 12 mM K+ to depolarize presynaptic neurons and glia produced a persistent increase in tonic current amplitude. The K+-induced increase in tonic current was reversibly inhibited by SKF89976a (40 μM), indicating that this was caused by nonvesicular GABA release from GABA transporter type 1 (GAT1). Nonvesicular GABA release due to GAT1 reversal also occurred in acute hippocampal brain slices. Our results indicate that tonic GABA currents are rapidly regulated by GABA-induced changes in intracellular Cl− concentration, PDP of extrasynaptic GABAA receptors, and nonvesicular GABA release. These mechanisms may influence tonic inhibition during seizures when neurons are robustly depolarized and extracellular GABA and K+ concentrations are elevated.

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Unique actions of GABA arising from cytoplasmic chloride microdomains

10.1101/2020.06.29.178160 ◽

2020 ◽

Author(s):

Negah Rahmati ◽

Kieran P. Normoyle ◽

Joseph Glykys ◽

Volodymyr I. Dzhala ◽

Kyle P. Lillis ◽

...

Keyword(s):

Fluorescent Protein ◽

Reversal Potential ◽

Receptor Activation ◽

Fluorescence Lifetime Imaging ◽

Lifetime Imaging ◽

Electrophysiological Measurements ◽

Highly Correlated ◽

Cation Chloride Cotransporters ◽

The Impact

AbstractDevelopmental, cellular, and subcellular variations in the direction of neuronal Cl− currents elicited by GABAA receptor activation have been frequently reported, and we found a corresponding variance in the reversal potential (EGABA) for individual interneurons synapsing on a single pyramidal cell. These findings suggest a corresponding variance in the cytoplasmic concentration of Cl− ([Cl−i]). We determined [Cl−]i by: 1) two-photon imaging of the Cl− sensitive, ratiometric fluorescent protein SuperClomeleon (sCLM); 2) Fluorescence Lifetime IMaging (FLIM) of the Cl− sensitive fluorophore MEQ; and 3) electrophysiological measurements of EGABA. These methods collectively demonstrated stable [Cl−]i microdomains in individual neurons in vivo. Fluorometric and electrophysiological estimates of local [Cl−]i were highly correlated. [Cl−]i microdomains persisted after pharmacological inhibition of cation-chloride cotransporters (CCCs) but steadily decreased after inhibiting the polymerization of the anionic macromolecule actin. These studies highlight the existence of functionally significant neuronal Cl− microdomains that modify the impact of GABAergic inputs.

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Inhibitory ultrapotent chemogenetics activate dopamine D1 receptor-expressing medium spiny neurons

10.1101/2020.07.01.181925 ◽

2020 ◽

Author(s):

Stephanie C. Gantz ◽

Maria M. Ortiz ◽

Andrew J. Belilos ◽

Khaled Moussawi

Keyword(s):

Brain Slices ◽

Reversal Potential ◽

Cell Types ◽

Receptor Activation ◽

Dopamine D1 Receptor ◽

D1 Receptor ◽

Medium Spiny Neurons ◽

Inhibitory Function ◽

Spiny Neurons

SUMMARYUltrapotent chemogenetics, including the chloride-permeable inhibitory PSAM4-GlyR receptor, were recently proposed as a powerful strategy to selectively control neuronal activity in awake, behaving animals. We aimed to validate the inhibitory function of PSAM4-GlyR in dopamine D1 receptor-expressing medium spiny neurons (D1-MSNs) in the ventral striatum. Activation of PSAM4-GlyR with the uPSEM792 ligand enhanced rather than suppressed the activity of D1-MSNs in vivo as indicated by increased c-fos expression in D1-MSNs. Whole-cell recordings in mouse brain slices showed that activation of PSAM4-GlyR did not inhibit firing of action potentials in D1-MSNs. Activation of PSAM4-GlyR depolarized D1-MSNs, attenuated GABAergic inhibition, and shifted the reversal potential of PSAM4-GlyR current to more depolarized potentials, perpetuating the depolarizing effect of receptor activation. The data show that ‘inhibitory’ PSAM4-GlyR chemogenetics may actually activate certain cell types, and highlight the pitfalls of utilizing chloride conductances to inhibit neurons.

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Reversible Loss of Hippocampal Function in a Mouse Model of Demyelination/Remyelination

10.1101/866723 ◽

2019 ◽

Author(s):

Aniruddha Das ◽

Chinthasagar Bastian ◽

Lexie Trestan ◽

Jason Suh ◽

Tanujit Dey ◽

...

Keyword(s):

Synaptic Transmission ◽

Hippocampal Neurons ◽

Brain Activity ◽

Neuronal Firing ◽

Demyelinating Diseases ◽

Therapeutic Interventions ◽

Electrophysiological Recordings ◽

Nonlinear Imaging

AbstractDemyelination of axons in the central nervous system (CNS) is a hallmark of multiple sclerosis (MS) and other demyelinating diseases. Cycles of demyelination, followed by remyelination, appear in the majority of MS patients, and are associated with the onset and quiescence of disease-related symptoms, respectively. Previous studies have shown in human patients and animal models that vast demyelination is accompanied by wide-scale changes to brain activity, but details of this process are poorly understood. We use electrophysiological recordings and nonlinear imaging of fluorescence from genetically-encoded calcium indicators to monitor the activity of hippocampal neurons during demyelination and remyelination processes over a period of 100 days. We find in vitro that synaptic transmission in CA1 neurons is diminished, and in vivo both CA1 and dentate gyrus (DG) neuronal firing rates are substantially reduced during demyelination and partially recover after a short remyelination period. This new approach allows monitoring how synaptic transmission changes, induced by cuprizone diet, are affecting neuronal activity, and can potentially be used to study the effects of therapeutic interventions in protecting the functionality of CNS neurons.

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