Abstract
3-Iodothyronamine (3-T1AM) is an endogenous thyroid hormone metabolite. The profound pharmacological effects of 3-T1AM on energy metabolism and thermal homeostasis have raised interest to elucidate its signaling properties in tissues that pertain to metabolic regulation and thermogenesis. Previous studies identified G protein-coupled receptors (GPCRs) and transient receptor potential channels (TRPs) as targets of 3-T1AM in different cell types. These two superfamilies of membrane proteins are largely expressed in tissue which influences energy balance and metabolism. As the first indication that 3-T1AM virtually modulates the function of the neurons in hypothalamus, we observed that intraperitoneal administration of 50 mg/kg bodyweight of 3-T1AM significantly increased the c-FOS activation in the paraventricular nucleus (PVN) of C57BL/6 mice. To elucidate the underlying mechanism behind this 3-T1AM-induced signalosome, we used three different murine hypothalamic cell lines, which are all known to express PVN markers, GT1-7, mHypoE-N39 (N39) and mHypoE-N41 (N41). Various aminergic GPCRs, which are the known targets of 3-T1AM, as well as numerous members of TRP channel superfamily, are expressed in these cell lines. Effects of 3-T1AM on activation of GPCRs were tested for the two major signaling pathways, the action of Gαs/adenylyl cyclase and Gi/o. Here, we demonstrated that this thyroid hormone metabolite has no significant effect on Gi/o signaling and only a minor effect on the Gαs/adenylyl cyclase pathway, despite the expression of known GPCR targets of 3-T1AM. Next, to test for other potential mechanisms involved in 3-T1AM-induced c-FOS activation in PVN, we evaluated the effect of 3-T1AM on the intracellular Ca2+ concentration and whole-cell currents. The fluorescence-optic measurements showed a significant increase of intracellular Ca2+ concentration in the three cell lines in the presence of 10 μM 3-T1AM. Furthermore, this thyroid hormone metabolite led to an increase of whole-cell currents in N41 cells. Interestingly, the TRPM8 selective inhibitor (10 μM AMTB) reduced the 3-T1AM stimulatory effects on cytosolic Ca2+ and whole-cell currents. Our results suggest that the profound pharmacological effects of 3-T1AM on selected brain nuclei of murine hypothalamus, which are known to be involved in energy metabolism and thermoregulation, might be partially attributable to TRP channel activation in hypothalamic cells.
Highlights
3-iodothyronamine (3-T1AM) is a decarboxylated and deiodinated derivative of thyroid hormones [1, 2]
To investigate whether 3-T1AM is capable of activating hypothalamic neurons in these nuclei in vivo, we performed intraperitoneal injections of 50 mg/kg bodyweight of 3-T1AM or dimethyl sulfoxide (DMSO)/PBS as control, six mice per group, and monitored 3-T1AM-induced neuron activation relative to DMSO/PBS
One hour after 3-T1AM injection, increased c-FOS staining of distinct neurons was clearly visible in the paraventricular nucleus (PVN) (60 ± 13 c-FOS positive cells per brain slide), while DMSO/PBS treated mice showed only few c-FOS positive neurons (16 ± 4 c-FOS positive cells per brain slide) (Figures 1A,B). 3-T1AM had no stimulatory effect on the medial preoptic area (MPO), the supraoptic nucleus (SON), the dorsolmedial nucleus of the hypothalamus, the periaqueductal gray (PAG) and the ventral tegmental segment (VTA) (Figure 1B and Supplemental Figure 1)
Summary
3-iodothyronamine (3-T1AM) is a decarboxylated and deiodinated derivative of thyroid hormones [1, 2]. Due to the profound pharmacological effects of 3T1AM and its accumulation in the selected tissue, numerous studies over the last years have been devoted to investigating the signaling property of this thyroid hormone metabolite. Different studies described several other GPCRs as 3-T1AM targets, mainly in vitro and in overexpressing systems. The same function principle applies to TRPV1, which is known as a heat- and capsacin receptor [25] Together, these properties implicate these TRPs as possible transducers of cold or warm stimuli within the hypothalamus [26], and in keratintocytes of human skin [27] and neurons on human corneal nerve fibers [28, 29]. The results of this study show a stimulatory effect of 3T1AM on Ca2+ mobilization and whole-cell currents in murine hypothalamic cells and that this effect is associated with TRPM8 activation
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