In this study, the gene expression of the α1, α2, α4, β1, β2, γ1, γ2, and δ GABAA receptor subunits was detected in THP-1 cells. The gene expression of α1, α4, β1, β2, γ1, and γ2 increased during macrophage differentiation. The gene expression of α1, α4, β1, β2, γ2, and δ decreased during M1 differentiation; additionally, propofol administration increased the expression of α1, α4, and β2 during M1 differentiation.
GABA is a major inhibitory neurotransmitter in the mammalian central nervous system. GABAA receptors, the primary target of GABA, are pentameric complexes consisting of three different subunits. Various combinations of GABAA receptor subunits determine receptor function. Mammals express 20-30 different GABAA receptor isoforms. The most common combination of GABAA receptor subunits consists of two α subunits, two β subunits, and one γ or δ subunit [18]. In this study, we investigated 13 GABAA receptor subunits: α1-6, β1-3, γ1-3, and δ. Most of the hypnotic effects of anesthetic agents are produced by the activation of GABAA receptors. Classical benzodiazepines, such as diazepam, bind to two distinct binding sites on the receptor. Benzodiazepines bind to the α-γ subunit interface present in the extracellular domain of the receptor. Benzodiazepines also bind to the β-α interface and γ-β interface, which are present in the transmembrane domain. Propofol binds to the β-α interface present in the transmembrane domain of the receptor [19].
GABAA receptors are present on immune cells, and GABAA signaling modifies immune function. GABAA receptors have been found on cluster of differentiation (CD)4+ and CD8+ T cells, macrophages, monocytes, and THP-1 cells [1–4]. The administration of diazepam in a mouse pneumonia model leads to immunosuppression, resulting in higher mortality, while the administration of the GABAA receptor antagonist, bicuculline, counteracts the immunosuppressive effect of diazepam and decreases mortality [20]. Another study reported that benzodiazepines are involved in immunosuppressive effect, resulting in increased mortality and the incidence of pneumonia [21]. The expression of GABAA receptor subunits on T cells and monocytes is modified by influenza infection, and diazepam administration affects immune function and increases susceptibility to infection [17]. These results indicate that the expression of GABAA receptor subunits can be modified by external stimuli, such as inflammation, differentiation, and drug administration, including the administration of intravenous anesthetics.
Our previous study showed that via the effects on GABAA receptors in THP-1 cells, propofol suppresses the production of inflammatory cytokines, IL-6 and IL-1β, during the differentiation into inflammatory M1 macrophage-like cells without affecting M1 differentiation [16]. The present study show that the gene expression of α1, α4, β2, and other subunits of GABAA receptors decreased during M1 differentiation, whereas addition of propofol increased the gene expression of α1, α4, and β2 subunits during M1 differentiation. Taken together, our previous and present data suggest that the suppression of inflammatory cytokines production during M1 differentiation in THP-1 cells by propofol administration may be associated with increasing the gene expression of α1, α4, and β2 subunits.
GABAA receptors have already been reported to be involved in inflammatory diseases, such as asthma, intestinal inflammation, and pulmonary fibrosis. The GABAA receptor, α1, is observed in human alveolar macrophages and monocytes and is responsible for diazepam-induced immunosuppression [17, 20]. When the GABAA receptor, α4, is knocked out in mice suffering from asthma, lung inflammation and airway reactivity deteriorate further [22]. In mice with stress-induced intestinal inflammation, α1, α4, and α5 GABAA receptor agonists exert anti-inflammatory effects, while α3 receptor agonists exacerbate inflammation [23]. Diazepam administration activates GABAA α4 receptors, thereby suppressing LPS-induced lung injury and the development of pulmonary fibrosis [24]. These results suggest that α1 and α4 GABAA receptors are involved in the suppression of inflammatory responses. In addition to such previous data, our data suggest that propofol suppresses production of inflammatiory cytokines in THP-1 cells during the M1 differentiation and propofol increases gene expressions of α1, α4, and β2 subunits of GABAA receptors during M1 differenctiation. Taken together, it is quite possible that propofol exerts its immunosuppressive effect by increasing expressions of α1 and α4 subunits of GABAa receptors.
This study has several limitations. First, an artificial THP-1 cell line was used; therefore, macrophages from human peripheral blood should be examined in a clinical setting in the next study, as the results from peripheral blood can be close to those acquired in actual situations and may easily be applied in clinical therapy. Second, only gene expression was analyzed in this study; therefore, changes should be examined at the protein level to clarify the precise mechanism of immunosuppressive effect via the GABAA receptor. Third, we analyzed only 13 major subunits; however, all 19 GABAA receptor subunits should be investigated in order to acquire more accurate results. Further studies are required to address these limitations.