GABA-A Alpha 2/3 but Not Alpha 1 Receptor Subunit Ligand Inhibits Harmaline and Pimozide-Induced Tremor in Rats.

Treatment of tremors, such as in essential tremor (ET) and Parkinson's disease (PD) is mostly ineffective. Exact tremor pathomechanisms are unknown and relevant animal models are missing. GABA-A receptor is a target for tremorolytic medications, but current non-selective drugs produce side effects and have safety liabilities. The aim of this study was a search for GABA-A subunit-specific tremorolytics using different tremor-generating mechanisms. Two selective positive allosteric modulators (PAMs) were tested. Zolpidem, targeting GABA-A α1, was not effective in models of harmaline-induced ET, pimozide- or tetrabenazine-induced tremulous jaw movements (TJMs), while the novel GABA-A α2/3 selective MP-III-024 significantly reduced both the harmaline-induced ET tremor and pimozide-induced TJMs. While zolpidem decreased the locomotor activity of the rats, MP-III-024 produced small increases. These results provide important new clues into tremor suppression mechanisms initiated by the enhancement of GABA-driven inhibition in pathways controlled by α2/3 but not α1 containing GABA-A receptors. Tremor suppression by MP-III-024 provides a compelling reason to consider selective PAMs targeting α2/3-containing GABA-A receptors as novel therapeutic drug targets for ET and PD-associated tremor. The possibility of the improved tolerability and safety of this mechanism over non-selective GABA potentiation provides an additional rationale to further pursue the selective α2/3 hypothesis.

Diet-induced obesity aggravates NK cell-mediated contact hypersensitivity reaction in Rag1-/- mice.

BACKGROUND: Previous studies showed that natural killer (NK) cells mediate contact hypersensitivity (CHS) reaction. Many reports are showing that obesity promotes several inflammatory diseases. It was shown that diet-induced obesity (DIO) aggravates classical T cell-mediated CHS in mice. OBJECTIVES: To determine whether the high-fat diet (HFD)-induced obesity modulates antigen-specific NK cell-mediated response. METHODS: We evaluated the effect of DIO on NK cell-mediated CHS reaction using a model of dinitrofluorobenzene (DNFB)-induced CHS in Rag1-/- mice. RESULTS: Rag1-/- mice fed HFD for 8 but not for 4 weeks developed aggravated CHS reaction determined by ear swelling measurement when compared to animals kept on normal diet (ND) prior to DNFB sensitization. The obese Rag1-/- mice presented the adipose tissue inflammation. Furthermore, in vitro analysis showed that feeding with HFD significantly increases interferon γ (IFN-γ) and interleukin (IL)-12p70 and decreases adiponectin concentration in liver mononuclear cell (LMNC) culture supernatants. The flow cytometry analysis of LMNC revealed that HFD treatment prior to DNFB sensitization increases the percentage of NK1.1+ IFN-γ+ cell population and affects the development and maturation of NK1.1+ cells. CONCLUSIONS: In summary, current results suggest that the DIO significantly modulates the local and systemic inflammatory response, contributing to exacerbation of the CHS response mediated by liver NK cells.

Cyclophosphamide-modified murine peritoneal macrophages induce CD4+ T contrasuppressor cells that protect contact sensitivity T effector cells from suppression.

BACKGROUND: Cyclophosphamide (CY) is one of the most widely used alkylating agents in the treatment of various cancers and some autoimmune diseases. Numerous reports suggest that CY exerts immunoregulatory effects. Animal studies have shown CY affects contact sensitivity (CS) response by depleting CD4+CD25+ T regulatory cells and CD8+ T suppressor (Ts) cells. In a mouse model of CS, we previously showed that in vivo treatment with CY shapes the immunogenic/immunoregulatory balance of peritoneal macrophages. The aim of the current study is to verify if macrophages (Mf) from CY-treated mice are indeed able to induce immunoregulatory cells that could protect from suppression. METHODS: Adoptive cell transfer of CS was used to examine immunomodulating properties of peritoneal Mf from CY-treated mice. Isolation of peritoneal Mf from animals that were (Mf-CY) or were not (Mf) treated with CY were cultured to identify cytokine repertoire. Further, we assessed spleen cell (SPLC) cytokine production following immunization with trinitrophenyl-conjugated Mf from donors treated (TNP-Mf-CY) or non-treated (TNP-Mf) with CY. RESULTS: In vitro experiments identified that Mf-CY produce more IL-6, TNF-α and TGF-ß than naïve Mf. Further, immunization with peritoneal TNP-Mf-CY induces CD4+ T contrasuppressor cells (Tcs) cells that protect CS-effector cells from suppression. Higher IL-17A secretion was observed from TNP-Mf-CY-treated mouse SPLC compared to SPLC from TNP-Mf injected mice suggesting that this cytokine might be important in mediating contrasuppression in this model. CONCLUSIONS: Our results show that in vivo treatment with CY influences mouse peritoneal Mf to induce CD4+ Tcs cells that protect CS-effector cells from suppressive signals of Ts cells.

Epicutaneous (EC) immunization with type II collagen (COLL II) induces CD4(+) CD8(+) T suppressor cells that protect from collagen-induced arthritis (CIA).

BACKGROUND: We have shown previously that epicutaneous (EC) immunization with protein antigen induces T suppressor cells that alleviate inflammatory response in contact hypersensitivity reactions, in an animal model of multiple sclerosis, and in TNBS-induced colitis. METHODS: DBA/1 mice were EC immunized with type II collagen (COLL II) spread over a gauze patch on days 0 and 4. On day 7, patches were removed and mice were intradermally (id) immunized with COLL II in CFA to induce collagen-induced arthritis (CIA). RESULTS: Our work shows that EC immunization with 100µg of COLL II prior to CIA induction reduces disease severity as determined by macroscopic evaluation. Reduced disease severity after EC immunization with COLL II correlates with milder histological changes found in joint sections. Experiments with the three non-cross-reacting antigens COLL II, ovalbumin (OVA) and myelin basic protein (MBP) showed that skin-induced suppression is antigen non-specific. Transfer experiments show that EC immunization with COLL II induces suppressor cells that belong to the population of CD4(+) CD8(+) double positive lymphocytes. Flow cytometry experiments showed increased percentage of CD4(+) CD8(+) RORγt(+) cells in axillary and inguinal lymph nodes isolated from mice patched with COLL II. CONCLUSION: Maneuver of EC immunization with a protein antigen that induces suppressor cells to inhibit inflammatory responses may become an attractive, noninvasive, needle-free therapeutic method for different clinical situations.