Epilepsy, Hyperalgesia, Impaired Memory, and Loss of Pre- and Postsynaptic GABAB Responses in Mice Lacking GABAB(1)

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Type B ␥-aminobutyric acid receptor (GABABR) is a G proteincoupled receptor that regulates neurotransmitter release and neuronal excitability throughout the brain. In various neurons, GABA BRs are concentrated at excitatory synapses. Although these receptors are assumed to respond to GABA spillover from neighboring inhibitory synapses, their function is not fully understood. Here we show a previously undescribed function of GABA BR exerted independent of GABA. In cerebellar Purkinje cells, interaction of GABA BR with extracellular Ca 2؉ (Ca o 2؉ ) leads to a constitutive increase in the glutamate sensitivity of metabotropic glutamate receptor 1 (mGluR1). mGluR1 sensitization is clearly mediated by GABA BR because it is absent in GABABR1 subunitknockout cells. However, the mGluR1 sensitization does not require G i/o proteins that mediate the GABABR's classical functions. Moreover, coimmunoprecipitation reveals complex formation between GABA BR and mGluR1 in the cerebellum. These findings demonstrate that GABA BR can act as Ca o 2؉ -dependent cofactors to enhance neuronal metabotropic glutamate signaling.T he type B ␥-aminobutyric acid receptor (GABA B R) is a G protein-coupled receptor (GPCR) distributed throughout the brain (1-3). GABA B R regulates neurotransmitter release and neuronal excitability via G i/o proteins (4). In the classic view, GABA B R responds to GABA released from inhibitory presynaptic terminals (4). However, in some central neurons including cerebellar Purkinje cells, postsynaptic GABA B Rs are concentrated perisynaptically at the excitatory synapses and present sparsely at the inhibitory synapses (5-7). Because GABA B Rs are insensitive to the excitatory neurotransmitter glutamate, a physiological role of GABA B R at excitatory synapses was assumed to depend on ␥-aminobutyric acid (GABA) spillover from neighboring inhibitory synapses (8, 9).The extracellular domain of GABA B R has an amino acid sequence homology to that of Ca 2ϩ -sensing receptor (CaR) (10). Some studies in the heterologous expression systems (11,12) revealed that GABA B R indeed interacts with extracellular Ca 2ϩ (Ca o 2ϩ ). Point mutation experiments indicate that the proximity of the GABA-binding site of GABA B R1 subunit (GBR1) is responsible for this interaction (11). Although Ca o 2ϩ -GABA B R interaction does not activate G proteins (12), it causes a remarkable conformational change of GABA B R as Ca o 2ϩ allosterically shifts GABA-GABA B R affinity (11,12 (22-25), and developmental synapse elimination (24, 26). In cerebellar Purkinje cells, mGluR1 colocalizes with GABA B R at the annuli of the dendritic spines innervated by excitatory parallel fibers (7,27). We have previously shown that mGluR1 signaling in Purkinje cells is enhanced as a consequence of interaction between Ca o 2ϩ and an unknown surface molecule(s) (28). For the reasons mentioned above, we considered GABA B R as a likely candidate for such a surface molecule.In Purkinje cells, mGluR1 outnumbers the other mGluR subtypes (16) and operates an inward ca...

Section: Discussionsupporting

confidence: 76%

Section: Absence Of the Ca O 2؉ -Dependent Mglur1 Sensitization In Gbmentioning

confidence: 99%

Ca ²⁺ activity at GABA _B receptors constitutively promotes metabotropic glutamate signaling in the absence of GABA

Tabata

Araishi

Hashimoto

et al. 2004

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“…The GABA B1 subunit is essential for the assembly of functional GABA B receptors and mice lacking the GABA B1 subunit show a complete absence of GABA B responses (17). To obtain mice with a deletion of the GABA B1 subunit gene restricted to orexinproducing neurons, orexin-Cre transgenic mice (SI Materials and Methods and Fig.…”

Section: Mice Selectively Lacking Functional Gabab Receptors In Oreximentioning

confidence: 99%

Selective loss of GABA _B receptors in orexin-producing neurons results in disrupted sleep/wakefulness architecture

Matsuki

Nomiyama

Takahira³

et al. 2009

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114

“…Moreover, in the absence of drug treatment these mice also express numerous pathologic behaviors such as epilepsy, hyperalgesia, and impaired memory (24,25). To circumvent these problems, which could be due to compensatory developmental responses of knockout mice, we planned our experiments such that RNAi was used for silencing of GABA B R1 subunits only in adult Drosophila.…”

mentioning

confidence: 99%

γ-Aminobutyric acid B receptor 1 mediates behavior-impairing actions of alcohol in Drosophila : Adult RNA interference and pharmacological evidence

Dzitoyeva

Dimitrijević

Manev

2003

111

In addition to their physiological function, metabotropic receptors for neurotransmitter ␥-aminobutyric acid (GABA), the GABAB receptors, may play a role in the behavioral actions of addictive compounds. Recently, GABAB receptors were cloned in fruit flies (Drosophila melanogaster), indicating that the advantages of this experimental model could be applied to GABAB receptor research. RNA interference (RNAi) is an endogenous process triggered by double-stranded RNA and is being used as a tool for functional gene silencing and functional genomics. Here we show how cell-nonautonomous RNAi can be induced in adult fruit flies to silence a subtype of GABAB receptors, GABABR1, and how RNAi combined with pharmacobehavioral techniques (including intraabdominal injections of active compounds and a computer-assisted quantification of behavior) can be used to functionally characterize these receptors. We observed that injection of double-stranded RNA complementary to GABABR1 into adult Drosophila selectively destroys GABABR1 mRNA and attenuates the behavioral actions of the GABAB agonist, 3-aminopropyl-(methyl)phosphinic acid. Moreover, both GABABR1 RNAi and the GABAB antagonist CGP 54626 reduced the behavior-impairing effects of ethanol, suggesting a putative role for the Drosophila GABAB receptors in alcohol's mechanism of action. The Drosophila model we have developed can be used for further in vivo functional characterization of GABAB receptor subunits and their involvement in the molecular and systemic actions of addictive substances.fruit fly ͉ GABAB receptors ͉ ethanol ͉ CGP 54626 ͉ 3-APMPA F ruit flies are used for research primarily because of the homology of Drosophila genes to those of mammals and because Drosophila is amenable to genetic manipulation including gene silencing through a process known as RNA interference (RNAi) (1-7). Typically, RNAi is triggered by double-stranded RNA (dsRNA), which is first processed by an RNase, Dicer (8), into 21-to 23-nt fragments. These fragments form a silencing complex that binds specifically to the dsRNA-complementary endogenous mRNA and leads to the destruction of the mRNA (1, 9). Injecting adult Drosophila intraabdominally with dsRNA results in the cell-nonautonomous silencing of the complementary endogenous mRNA throughout the body, including the CNS (10). Because this method does not interfere in the normal development of the animal but can replicate typical phenotypes produced by gene mutations (11), here we have used adult RNAi to silence ␥-aminobutyric acid (GABA) B receptors.The slow inhibitory GABA synaptic neurotransmission is mediated by the metabotropic G protein-coupled and cAMPlinked GABA B receptors (12, 13) that have been cloned in humans (14) and also recently in Drosophila (15). In addition to their physiological function, the GABA B receptors may play a role in the behavioral actions of addictive compounds such as ethanol (16)(17)(18)(19). Although Drosophila has been used to study the mechanisms of action of alcohol (20)(21)(22)(23), as yet, this model ...