Teemu Aitta-aho

Reduced aggression in AMPA-type glutamate receptor GluR-A subunit-deficient mice

The importance of AMPA‐type glutamate receptors has been demonstrated in neuronal plasticity and in adaptation to drugs of abuse. We studied the involvement of AMPA receptors in social interaction and anxiety and found that in several paradigms of agonistic behavior naïve male mice deficient for the GluR‐A subunit‐ containing AMPA receptors are less aggressive than wild‐type littermates. GluR‐A deficient mice and wild‐type littermates exhibited similar basic behavior and reflexes as monitored by observational Irwins test, but they tended to be less anxious in elevated plus‐maze and light‐dark tests. Maternal aggression or male‐female encounters were not affected which suggests that male hormones are involved in the expression of suppressed aggressiveness. However, testosterone levels and brain monoamines can be excluded and found to be similar between GluR‐A deficient and wild‐type littermates. The reduced AMPA receptor levels caused by the lack of the GluR‐A subunit, and measured by a 30% reduction in hippocampal [3H]‐S‐AMPA binding, seem to be the reason for suppressed male aggressiveness. When we analyzed mice with reduced number of functional AMPA receptors mediated by the genomic introduced GluR‐A(Q582R) channel mutation, we observed again male‐specific suppressed aggression, providing additional evidence for GluR‐A subunit‐containing AMPA receptor involvement in aggression.

The in Vivo Contributions of TASK-1-Containing Channels to the Actions of Inhalation Anesthetics, the alpha~2 Adrenergic Sedative Dexmedetomidine, and Cannabinoid Agonists

Inhalation anesthetics activate and cannabinoid agonists inhibit TWIK-related acid-sensitive K+ channels (TASK)-1 two-pore domain leak K+ channels in vitro. Many neuromodulators, such as noradrenaline, might also manifest some of their actions by modifying TASK channel activity. Here, we have characterized the basal behavioral phenotype of TASK-1 knockout mice and tested their sensitivity to the inhalation anesthetics halothane and isoflurane, the α2 adrenoreceptor agonist dexmedetomidine, and the cannabinoid agonist WIN55212-2 mesylate [R-(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3,-de]-1,4-benzoxazinyl]-(1-naphtalenyl)methanone mesylate)]. TASK-1 knockout mice had a largely normal behavioral phenotype. Male, but not female, knockout mice displayed an enhanced acoustic startle response. The knockout mice showed increased sensitivity to thermal nociception in a hot-plate test but not in a tail-flick test. The analgesic, sedative, and hypothermic effects of WIN55212-2 (2–6 mg/kg s.c.) were reduced in TASK-1 knockout mice. These results implicate TASK-1-containing channels in supraspinal pain pathways, in particular those modulated by endogenous cannabinoids. TASK-1 knockout mice were less sensitive to the anesthetic effects of halothane and isoflurane than wild-type littermates, requiring higher anesthetic concentrations to induce immobility as reflected by loss of the tail-withdrawal reflex. Our results support the idea that the activation of multiple background K+ channels is crucial for the high potency of inhalation anesthetics. Furthermore, TASK-1 knockout mice were less sensitive to the sedative effects of dexmedetomidine (0.03 mg/kg s.c.), suggesting a role for the TASK-1 channels in the modulation of function of the adrenergic locus coeruleus nuclei and/or other neuronal systems.

Optogenetic Evidence for Inhibitory Signaling from Orexin to MCH Neurons via Local Microcircuits

The lateral hypothalamus (LH) is a key regulator of multiple vital behaviors. The firing of brain-wide-projecting LH neurons releases neuropeptides promoting wakefulness (orexin/hypocretin; OH), or sleep (melanin-concentrating hormone; MCH). OH neurons, which coexpress glutamate and dynorphin, have been proposed to excite their neighbors, including MCH neurons, suggesting that LH may sometimes coengage its antagonistic outputs. However, it remains unclear if, when, and how OH actions promote temporal separation of the sleep and wake signals, a process that fails in narcolepsy caused by OH loss. To explore this directly, we paired optogenetic stimulation of OH cells (at rates that promoted awakening in vivo) with electrical monitoring of MCH cells in mouse brain slices. Membrane potential recordings showed that OH cell firing inhibited action potential firing in most MCH neurons, an effect that required GABAA but not dynorphin receptors. Membrane current analysis showed that OH cell firing increased the frequency of fast GABAergic currents in MCH cells, an effect blocked by antagonists of OH but not dynorphin or glutamate receptors, and mimicked by bath-applied OH peptide. In turn, neural network imaging with a calcium indicator genetically targeted to MCH neurons showed that excitation by bath-applied OH peptides occurs in a minority of MCH cells. Collectively, our data provide functional microcircuit evidence that intra-LH feedforward loops may facilitate appropriate switching between sleep and wake signals, potentially preventing sleep disorders.

Histamine and H3 receptor-dependent mechanisms regulate ethanol stimulation and conditioned place preference in mice

RationaleNeuronal histamine has a prominent role in sleep–wake control and body homeostasis, but a number of studies suggest that histamine has also a role in higher brain functions including drug reward.ObjectiveThe present experiments characterized the involvement of histamine and its H3 receptor in ethanol-related behaviors in mice.Materials and methodsMale histidine decarboxylase knockout (HDC KO) and control mice were used to study the role of histamine in ethanol-induced stimulation of locomotor activity, impairment of motor coordination, and conditioned place preference (CPP). Male C57BL/6Sca mice were used to study the effects of H3 receptor antagonist in the effects of ethanol on locomotor activity.ResultsThe HDC KO mice displayed a weaker stimulatory response to acute ethanol than the wild-type (WT) mice. No differences between genotypes were found after ethanol administration on accelerating rotarod. The HDC KO mice showed stronger ethanol-induced CPP than the WT mice. Binding of the GABAA receptor ligand [3H]Ro15-4513 was not markedly changed in HDC KO mouse brain and thus could not explain altered responses in KO mice. Ethanol increased the activity of C57BL/6Sca mice, and H3 receptor antagonist ciproxifan inhibited this stimulation. In CPP paradigm ciproxifan, an H3 receptor inverse agonist potentiated ethanol reward.ConclusionsHistaminergic neurotransmission seems to be necessary for the stimulatory effect of ethanol to occur, whereas lack of histamine leads to changes that enhance the conditioned reward by ethanol. Our findings also suggest a role for histamine H3 receptor in modulation of the ethanol stimulation and reward.

Acute effects of AMPA-type glutamate receptor antagonists on intermale social behavior in two mouse lines bidirectionally selected for offensive aggression

Involvement of AMPA-type glutamate receptors in the regulation of social behavior has been suggested by experiments with mice deficient for the GluR-A subunit-containing AMPA receptors showing reduced intermale aggression. In the present study, effects of AMPA receptor antagonists on mouse social behavior towards unfamiliar Swiss-Webster males on a neutral territory were tested using male subjects from the Turku Aggressive (TA) and Turku Non-Aggressive (TNA) mouse lines bidirectionally selected for high and low levels of offensive aggression. The drugs were the competitive antagonists 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide (NBQX), and the non-competitive antagonist 4-(8-methyl-9H-1,3-dioxolo[4,5-h][2,3]benzodiazepin-5-yl)-benzenamine (GYKI 52466). In TA mice, CNQX and NBQX decreased the biting component of aggressive structure, while GYKI 52466 suppressed all aggressive manifestations. All drugs increased anxiety-like behavior towards the partner. In TNA mice, NBQX activated mouse social behavior and ambivalent aggression, while CNQX and GYKI 52466 only increased anxiety. Thus, AMPA receptor antagonists affect aggressive behaviors in TA mice supporting the idea that AMPA receptors are involved in the modulation of agonistic impulsive behavioral pattern. GYKI 52466 appeared to be the most selective and efficacious in suppressing the aggression.

GABA Site Agonist Gaboxadol Induces Addiction-Predicting Persistent Changes in Ventral Tegmental Area Dopamine Neurons But Is Not Rewarding in Mice or Baboons

Dopamine neurons of the ventral tegmental area (VTA) are involved at early phases of drug addiction. Even the first in vivo dose of various abused drugs induces glutamate receptor plasticity at the excitatory synapses of these neurons. Benzodiazepines that suppress the inhibitory GABAergic interneurons in the VTA via facilitation of synaptic GABAA receptors have induced neuroplasticity in dopamine neurons due to this disinhibitory mechanism. Here, we have tested a non-benzodiazepine direct GABA site agonist 4,5,6,7-tetrahydroisoxazolol[4,5-c]pyridine-3-ol (THIP) (also known as gaboxadol) that acts preferentially via high-affinity extrasynaptic GABAA receptors. A single sedative dose of THIP (6 mg/kg) to mice induced glutamate receptor plasticity for at least 6 d after administration. Increased AMPA/NMDA receptor current ratio and increased frequency, amplitude, and rectification of AMPA receptor responses suggested persistent targeting of GluA2-lacking AMPA receptors in excitatory synapses of VTA dopamine neurons ex vivo after THIP administration. This effect was abolished in GABAA receptor δ−/− mice, which have a loss of extrasynaptic GABAA receptors. In behavioral experiments, we found neither acute reinforcement in intravenous self-administration sessions with THIP at relevant doses using a yoked control paradigm in mice nor in baboons using a standard paradigm for assessing drug abuse liability; nor was any place preference found after conditioning sessions with various doses of THIP but rather a persistent aversion in 6 mg/kg THIP-conditioned mice. In summary, we found that activation of extrasynaptic δ-subunit-containing GABAA receptors leads to glutamate receptor plasticity of VTA dopamine neurons, but is not rewarding, and, instead, induces aversion.

Excessive novelty-induced c-Fos expression and altered neurogenesis in the hippocampus of GluA1 knockout mice.

α‐Amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA) receptor GluA1 subunit‐deficient (GluA1−/−) mice display novelty‐induced hyperactivity, cognitive and social defects and may model psychiatric disorders, such as schizophrenia and depression/mania. We used c‐Fos expression in GluA1−/− mice to identify brain regions responsible for novelty‐induced hyperlocomotion. Exposure to a novel cage for 2 h significantly increased c‐Fos expression in many brain regions in both wild‐type and knockout mice. Interestingly, the clearest genotype effect was observed in the hippocampus and its main input region, the entorhinal cortex, where the novelty‐induced c‐Fos expression was more strongly enhanced in GluA1−/− mice. Their novelty‐induced hyperlocomotion partly depended on the activity of AMPA receptors, as it was diminished by the AMPA receptor antagonist 2,3‐dioxo‐6‐nitro‐1,2,3,4‐tetrahydrobenzo[f]quinoxaline‐7‐sulphonamide (NBQX) and unaffected by the AMPA receptor potentiator 2,3‐dihydro‐1,4‐benzodioxin‐6‐yl‐1‐piperidinylmethanone (CX546). The hyperlocomotion of GluA1−/− mice was normalised to the level of wild‐type mice within 5–6 h, after which their locomotion followed normal circadian rhythm and was not affected by acute or chronic treatments with the selective serotonin reuptake inhibitor escitalopram. We propose that hippocampal dysfunction, as evidenced by the excessive c‐Fos response to novelty, is the major contributor to novelty‐induced hyperlocomotion in GluA1−/− mice. Hippocampal dysfunction was also indicated by changes in proliferation and survival of adult‐born dentate gyrus cells in the knockout mice. These results suggest focusing on the functions of hippocampal formation, such as novelty detection, when using the GluA1−/− mouse line as a model for neuropsychiatric and cognitive disorders.

Neurosteroid Agonist at GABAA Receptor Induces Persistent Neuroplasticity in VTA Dopamine Neurons

The main fast-acting inhibitory receptors in the mammalian brain are γ-aminobutyric acid type-A (GABAA) receptors for which neurosteroids, a subclass of steroids synthesized de novo in the brain, constitute a group of endogenous ligands with the most potent positive modulatory actions known. Neurosteroids can act on all subtypes of GABAA receptors, with a preference for δ-subunit-containing receptors that mediate extrasynaptic tonic inhibition. Pathological conditions characterized by emotional and motivational disturbances are often associated with perturbation in the levels of endogenous neurosteroids. We studied the effects of ganaxolone (GAN)—a synthetic analog of endogenous allopregnanolone that lacks activity on nuclear steroid receptors—on the mesolimbic dopamine (DA) system involved in emotions and motivation. A single dose of GAN in young mice induced a dose-dependent, long-lasting neuroplasticity of glutamate synapses of DA neurons ex vivo in the ventral tegmental area (VTA). Increased α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)/N-methyl-D-aspartate ratio and rectification of AMPA receptor responses even at 6 days after GAN administration suggested persistent synaptic targeting of GluA2-lacking AMPA receptors. This glutamate neuroplasticity was not observed in GABAA receptor δ-subunit-knockout (δ-KO) mice. GAN (500 nM) applied locally to VTA selectively increased tonic inhibition of GABA interneurons and triggered potentiation of DA neurons within 4 h in vitro. Place-conditioning experiments in adult wild-type C57BL/6J and δ-KO mice revealed aversive properties of repeated GAN administration that were dependent on the δ-subunits. Prolonged neuroadaptation to neurosteroids in the VTA might contribute to both the physiology and pathophysiology underlying processes and changes in motivation, mood, cognition, and drug addiction.

Importance of GluA1 Subunit-Containing AMPA Glutamate Receptors for Morphine State-Dependency

In state-dependency, information retrieval is most efficient when the animal is in the same state as it was during the information acquisition. State-dependency has been implicated in a variety of learning and memory processes, but its mechanisms remain to be resolved. Here, mice deficient in AMPA-type glutamate receptor GluA1 subunits were first conditioned to morphine (10 or 20 mg/kg s.c. during eight sessions over four days) using an unbiased procedure, followed by testing for conditioned place preference at morphine states that were the same as or different from the one the mice were conditioned to. In GluA1 wildtype littermate mice the same-state morphine dose produced the greatest expression of place preference, while in the knockout mice no place preference was then detected. Both wildtype and knockout mice expressed moderate morphine-induced place preference when not at the morphine state (saline treatment at the test); in this case, place preference was weaker than that in the same-state test in wildtype mice. No correlation between place preference scores and locomotor activity during testing was found. Additionally, as compared to the controls, the knockout mice showed unchanged sensitization to morphine, morphine drug discrimination and brain regional μ-opioid receptor signal transduction at the G-protein level. However, the knockout mice failed to show increased AMPA/NMDA receptor current ratios in the ventral tegmental area dopamine neurons of midbrain slices after a single injection of morphine (10 mg/kg, s.c., sliced prepared 24 h afterwards), in contrast to the wildtype mice. The results indicate impaired drug-induced state-dependency in GluA1 knockout mice, correlating with impaired opioid-induced glutamate receptor neuroplasticity.

Reversal of novelty-induced hyperlocomotion and hippocampal c-Fos expression in GluA1 knockout male mice by the mGluR2/3 agonist LY354740

Dysfunctional glutamatergic neurotransmission has been implicated in schizophrenia and mood disorders. As a putative model for these disorders, a mouse line lacking the GluA1 subunit (GluA1-KO) of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) glutamate receptor displays a robust novelty-induced hyperlocomotion associated with excessive neuronal activation in the hippocampus. Agonists of metabotropic glutamate 2/3 receptors (mGluR2/3) inhibit glutamate release in various brain regions and they have been shown to inhibit neuronal activation in the hippocampus. Here, we tested a hypothesis that novelty-induced hyperlocomotion in the GluA1-KO mice is mediated via excessive hippocampal neuronal activation by analyzing whether an mGluR2/3 agonist inhibits this phenotypic feature. GluA1-KO mice and littermate wildtype (WT) controls were administered with (1S,2S,5R,6S)-2-aminobicyclo[3.1.0]hexane-2,6-dicarboxylic acid (LY354740) (15 mg/kg, i.p.) 30 min before a 2-h exposure to novel arenas after which c-Fos immunopositive cells were analyzed in the hippocampus. LY354740 (15 mg/kg) decreased hyperactivity in male GluA1-KO mice, with only a minimal effect in WT controls. This was observed in two cohorts of animals, one naïve to handling and injections, another pre-handled and accustomed to injections. LY354740 (15 mg/kg) also reduced the excessive c-Fos expression in the dorsal hippocampal CA1 pyramidal cell layer in maleGluA1-KO mice, while not affecting c-Fos levels in WT mice. In female mice, no significant effect for LY354740 (15 mg/kg) on hyperactive behavior or hippocampal c-Fos was observed in either genotype or treatment cohort. A higher dose of LY354740 (30 mg/kg) alleviated hyperlocomotion of GluA1-KO males, but not that of GluA1-KO females. In conclusion, the excessive behavioral hyperactivity of GluA1-KO mice can be partly prevented by reducing neuronal excitability in the hippocampus with the mGluR2/3 agonist suggesting that the hippocampal reactivity is strongly involved in the behavioral phenotype of GluA1-KO mice.