Mice Lacking GABA(A) Receptor delta Subunit Have Altered Pharmaco-EEG Responses to Multiple Drugs

In the brain, extrasynaptically expressed ionotropic, delta subunit-containing gamma-aminobutyric acid A-type receptors (delta-GABA(A)Rs) have been implicated in drug effects at both neuronal and behavioral levels. These alterations are supposed to be caused via drug-induced modulation of receptor ionophores affecting chloride ion-mediated inhibitory tonic currents. Often, a transgenic mouse model genetically lacking the delta-GABA(A)Rs (delta-KO) has been used to study the roles of delta-GABA(A)Rs in brain functions, because a specific antagonist of the delta-GABA(A)Rs is still lacking. We have previously observed with these delta-KO mice that activation of delta-GABA(A)Rs is needed for morphine-induced conditioning of place preference, and others have suggested that delta-GABA(A)Rs act as targets selectively for low doses of ethanol. Furthermore, activation of these receptors via drug-mediated agonism induces a robust increase in the slow-wave frequency bands of electroencephalography (EEG). Here, we tested delta-KO mice (compared to littermate wild-type controls) for the pharmaco-EEG responses of a broad spectrum of pharmacologically different drug classes, including alcohol, opioids, stimulants, and psychedelics. Gaboxadol (THIP), a known superagonist of delta-GABA(A)Rs, was included as the positive control, and as expected, delta-KO mice produced a blunted pharmaco-EEG response to 6 mg/kg THIP. Pharmaco-EEGs showed notable differences between treatments but also differences between delta-KO mice and their wild-type littermates. Interestingly mephedrone (4-MMC, 5 mg/kg), an amphetamine-like stimulant, had reduced effects in the delta-KO mice. The responses to ethanol (1 g/kg), LSD (0.2 mg/kg), and morphine (20 mg/kg) were similar in delta-KO and wild-type mice. Since stimulants are not known to act on delta-GABA(A)Rs, our findings on pharmaco-EEG effects of 4-MMC suggest that delta-GABA(A)Rs are involved in the secondary indirect regulation of the brain rhythms after 4-MMC.Peer reviewe

Increased Sensitivity of Mice Lacking Extrasynaptic delta-Containing GABA(A) Receptors to Histamine Receptor 3 Antagonists

Histamine/gamma-aminobutyric acid (GABA) neurons of posterior hypothalamus send wide projections to many brain areas and participate in stabilizing the wake state. Recent research has suggested that GABA released from the histamine/GABA neurons acts on extrasynaptic GABA(A) receptors and balances the excitatory effect of histamine. In the current study, we show the presence of vesicular GABA transporter mRNA in a majority of quantified hypothalamic histaminergic neurons, which suggest vesicular release of GABA. As histamine/GABA neurons form conventional synapses infrequently, it is possible that GABA released from these neurons diffuses to target areas by volume transmission and acts on extrasynaptic GABA receptors. To investigate this hypothesis, mice lacking extrasynaptic GABA(A) receptor delta subunit (Gabrd KO) were used. A pharmacological approach was employed to activate histamine/GABA neurons and induce histamine and presumably, GABA, release. Control and Gabrd KO mice were treated with histamine receptor 3 (Hrh3) inverse agonists ciproxifan and pitolisant, which block Hrh3 autoreceptors on histamine/GABA neurons and histamine-dependently promote wakefulness. Low doses of ciproxifan (1 mg/kg) and pitolisant (5 mg/kg) reduced locomotion in Gabrd KO, but not in WT mice. EEG recording showed that Gabrd KO mice were also more sensitive to the wake-promoting effect of ciproxifan (3 mg/kg) than control mice. Low frequency delta waves, associated with NREM sleep, were significantly suppressed in Gabrd KO mice compared with the WT group. Ciproxifan-induced wakefulness was blocked by histamine synthesis inhibitor alpha-fluoromethylhistidine (alpha FMH). The findings indicate that both histamine and GABA, released from histamine/GABA neurons, are involved in regulation of brain arousal states and delta-containing subunit GABA(A) receptors are involved in mediating GABA response.Peer reviewe

Alcohol Co-Administration Changes Mephedrone-Induced Alterations of Neuronal Activity

Mephedrone (4-MMC), despite its illegal status, is still a widely used psychoactive substance. Its effects closely mimic those of the classical stimulant drug methamphetamine (METH). Recent research suggests that unlike METH, 4-MMC is not neurotoxic on its own. However, the neurotoxic effects of 4-MMC may be precipitated under certain circumstances, such as administration at high ambient temperatures. Common use of 4-MMC in conjunction with alcohol raises the question whether this co-consumption could also precipitate neurotoxicity. A total of six groups of adolescent rats were treated twice daily for four consecutive days with vehicle, METH (5 mg/kg) or 4-MMC (30 mg/kg), with or without ethanol (1.5 g/kg). To investigate persistent delayed effects of the administrations at two weeks after the final treatments, manganese-enhanced magnetic resonance imaging brain scans were performed. Following the scans, brains were collected for Golgi staining and spine analysis. 4-MMC alone had only subtle effects on neuronal activity. When administered with ethanol, it produced a widespread pattern of deactivation, similar to what was seen with METH-treated rats. These effects were most profound in brain regions which are known to have high dopamine and serotonin activities including hippocampus, nucleus accumbens and caudate-putamen. In the regions showing the strongest activation changes, no morphological changes were observed in spine analysis. By itself 4-MMC showed few long-term effects. However, when co-administered with ethanol, the apparent functional adaptations were profound and comparable to those of neurotoxic METH.Peer reviewe

Neurotoxic and Extrasynaptic GABAA Receptor-Mediated Effects of Drugs in Rodent Models

Abstract Introduction | Drug usage is an increasing problem in the modern age affecting both society and individual users. In many circumstances, drugs are not used only one by one, but many compounds are often co-consumed. This co-consumption makes it relatively hard to predict the effects as multiple neurotransmitter systems are involved. Furthermore, different compounds can have pharmacodynamic and -kinetic interactions. This thesis focuses on studying how extrasynaptic GABAA receptors mediating tonic inhibition affect the effects of a broad spectrum of different drugs. Lastly, this thesis focuses on evaluating how ethanol affects the neurotoxic effects of stimulants. Methods | A transgenic mouse model, lacking δ subunit-containing extrasynaptic GABAA receptors, was compared to wild-type mice to reveal the actions of those receptors. These mice were used in multiple research paradigms ranging from electroencephalography to multiple behavioral tests. Manganese-enhanced magnetic resonance imaging and Golgi staining were used to assess the neurotoxicity of ethanol-stimulant co-consumption in rats. Results | It was observed that the reduced tonic inhibition in the mouse model increased opioid-induced antinociception but decreased the induced reward. However, the opioid-induced electroencephalography was not affected. Methamphetamine-like stimulant mephedrone induced only minor activations, whereas methamphetamine produced significant deactivations. However, co-consumption of ethanol made mephedrone-induced alterations similar to methamphetamine. Conclusions | The modulation of extrasynaptic GABAA receptors could have a beneficial role in the modulation of opioid reward and nociception. This thesis also suggests that mephedrone should not be considered as neurotoxic as methamphetamine, at least in normal ambient temperatures.Tiivistelmä Johdanto | Päihteiden yhteiskunnalliset ja yksilötasoiset haitat ovat tunnettu kasvava ongelma. Useissa tilanteissa päihteiden käyttö sisältää useiden erilaisten aineiden käyttämistä yhtä aikaa, jolloin lopullisen vaikutuksen tietäminen yhteisvaikutusten takia voi olla hyvinkin hankalaa. Tämä väitöskirja on keskittynyt tutkimaan, kuinka toonista estoa välittävät ekstrasynaptiset GABAA-reseptorit vaikuttavat useiden laajalti käytössä olevien huumausaineiden vaikutuksiin. Lisäksi väitöskirjassa tutkittiin alkoholin vaikutusta stimulanttien tuottamaan neurotoksisuuteen. Menetelmät | Ekstrasynaptisten GABAA-reseptorien tutkimiseen käytettiin transgeenistä hiirimallia, jolta puuttuvat δ-alayksikön sisältävät ekstrasynaptiset GABAA-reseptorit. Näitä hiiriä käytettiin tutkimusasetelmissa, joissa tutkittiin niiden vasteita useille eri aineille. Lisäksi stimulanttien neurotoksisuuden tutkimiseksi käytettiin mangaanitehostettua magneettikuvausta ja Golgi-värjäystä rotilla. Tulokset | Toonisen ekstrasynaptisen GABAA-reseptorivälitteisen inhibition väheneminen lisäsi opioidien kivunlievityskykyä, mutta vähensi niiden palkitsevaa vaikutusta. Opioidien tuottamaan aivosähkökäyrään ekstrasynaptisella inhibitiolla ei ollut kuitenkaan vaikutusta. Metamfetamiinin kaltaisen stimulantin mefedronin tuottamat toiminnalliset muutokset olivat huomattavasti vähäisempiä kuin metamfetamiinin, mutta etanolin samanaikainen annostelu muutti mefedronin vaikutukset metamfetamiinin kaltaisiksi. Mefedroni tai metamfetamiini eivät kuitenkaan tuottaneet rakenteellisia muutoksia aivoihin. Johtopäätökset | Ekstrasynaptisen GABAA-reseptorien modulaatiolla voisi olla rooli erityisesti opioidien palkitsevuuden vähentämisessä ja kivunlievityksen tehostamisessa. Lisäksi tutkimus tukee aiempaa käsitystä siitä, että mefedronia ei voida yksin pitää erityisen neurotoksisena