**2. Mode-of-action of TH in modulating metabolic efficiency**

The first description of TH-induced calorigenesis dates to 1895 (1). That initial report has been followed by exhaustive data focusing on the phenomenology of TH action as reflected by hyper- and hypothyroidsm. Thus, high levels of TH in mammals increase oxygen consumption and heat production, resulting in pronounced body weight loss, while low levels of TH are associated with a decrease in metabolic rate and the oxidation of energy substrates (glucose, fatty acids and amino acids), resulting in pronounced increase in body weight (2-6). Although it was widely accepted that TH stimulates calorigenesis by affecting respiration, its cellular mode-of-action remained to be resolved. Hence, exhaustive attempts were made by the scientific community throughout the twentieth century to verify the mechanism(s) involved in modulating metabolic efficiency by TH. Studies by Lardy and Feldott (7) and Hess and Martius (8) have pointed out during the 1950s, that the respiratory control ratio of isolated mitochondria was robustly decreased in the presence of added T4. TH was thus claimed to have direct action at the mitochondrial level by inducing 'mitochondrial uncoupling', namely, dissociating mitochondrial phosphorylation from its substrate oxidation driver. However, the high T4 doses used in those studies implied possible non-physiological activity rather than authentic TH-induced calorigenesis. Later

© 2012 Bar-Tana et al., licensee InTech. This is an open access chapter distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2012 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

evidence in support of 'mitochondrial uncoupling' has however indicated increase in oxygen consumption and mitochondrial proton permeability of isolated liver mitochondria derived from hyperthyroid rats, with concomitant decrease in mitochondrial phosphate potential and inner mitochondrial membrane (IMM) potential (9, 10). These observations were further corroborated by increase in liver oxidizing capacity of hyperthyroid rats accompanied by decrease in phosphate and cytosolic redox potential (11), while opposite effects were reported in livers of hypothyroid rats (12). Similarly, hepatocytes isolated from T3-treated rats show higher oxygen consumption and lower IMM potential as compared with non-treated control (13-16). Also, a decrease in IMM potential has been reported in TH-treated human lymphocytes or those derived from hyperthyroid patients (17). Overall, these findings suggested that TH indeed induces mitochondrial uncoupling, and that mitochondrial uncoupling may account for the cellular mode-of-action of TH in modulating metabolic efficiency *in vivo*.

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nucleotide translocase (ANT), cytochrome c1) (24, 25), or genes coding for intermediate factors that are indirectly involved in promoting the nuclear expression of mitochondrial components (i.e. nuclear respiratory factors 1 and -2, peroxisome proliferator-activated receptor-γ coactivator-1) (26), or in stimulating mitochondrial DNA replication (27). Hence, TH-induced mitochondrial uncoupling was believed to be accounted for by TH-induced gene expression of respective protein targets that modulate mitochondrial oxidative phosphorylation. In pursuing putative proteins involved in TH-induced mitochondrial uncoupling, a number of candidates have been suggested. These included the adenine nucleotide translocase, proteins that are involved in phosphatidylglycerol and cardiolipin

With the discovery of UCPs, extensive efforts were invested in verifying their putative role in mediating the calorigenic effect of TH. In fact, the UCP-coding genes have TREs in their promoters and their expression level is increased by TH treatment, implying their putative role in mediating TH-induced calorigenesis (40). UCP1 (29) mediates proton leak in brown adipose tissue IMM (30), resulting in uncoupling fuel oxidation from ATP synthesis and in dissipating IMM potential as heat. The adaptive thermogenic response of UCP1 is driven by the sympathetic nervous system in response to cold temperature or high-energy cafeteria diet, and could apparently serve as target for TH in modulating total body energy expenditure. Indeed, recent findings by Lopez et al (31) have indicated that TH treatment results in suppressing hypothalamic AMP-activated protein kinase (AMPK) activity, resulting in SNS-induced thermogenic response of brown adipose fat. However, UCP1 is specifically expressed in brown adipose tissue, which is sparse in adult humans. While recent findings point to some brown adipose islets in adult humans (32-37), their putative impact on total body energy expenditure still remains to be resolved. Hence, other proteins that share sequence homology with UCP1(38), including the ubiquitously expressed UCP2, and in particular the UCP3 that is expressed in skeletal muscle, were pursued for their role in mediating TH-induced calorigenesis (39). However, the following observations may indicate that UCP2 and UCP3 may not account for TH-induced mitochondrial uncoupling (41). Thus, findings suggest that UCP2/3 do not contribute to adaptive thermogenesis (42), but may have a role in ROS signaling (43) and/or in exporting fatty acid anions from the mitochondrial matrix (44). Also, the expression of liver UCP2/3 proteins is restricted to Kupffer cells, implying that the uncoupling effect of TH in liver parenchymal cells is not due to UCPs. Most importantly, UCP3 knock-out mice are lean and show normal response to TH (45), leaving unresolved the

synthesis (28), and in particular the mitochondrial uncoupling proteins (UCPs).

specific proteins that may mediate TH metabolic effects in the mitochondrial context.

In analogy to UCPs, mitochondria consist of Permeability Transition Pores (PTP) (46-50) located at the contact sites of the inner (IMM) and outer (OMM) mitochondrial membranes. The molecular composition and structure of mitochondrial PTP still remains to be resolved. The current model of PTP consists of the integral proteins ANT (in the IMM), the voltage-

**5. Mitochondrial permeability transition pore (PTP)** 

**4. Mitochondrial uncoupling proteins (UCPs)** 

Concomitantly with the proposed mitochondrial paradigm of TH, others have proposed non-mitochondrial activity of TH in modulating metabolic efficiency. Thus, TH was claimed to induce "futile substrate cycles", namely, opposing energy-requiring metabolic pathways that proceed simultaneously without generating net products, e.g. glycolysis accompanied by gluconeogenesis, lipolysis with lipogenesis (18, 19), Na+/K+ ATPase with concomitant Na+/K+ leakage (13), or glycerol-3- phosphate/NADH mitochondrial shuttling (20). However, these proposed mechanisms could account for only a small fraction (about 15%) of the total increase in oxygen consumption induced by TH (13, 21), resulting in a mitochondrial paradigm consensus for TH-induced calorigenesis. Yet, the concerned mitochondrial targets still remained enigmatic.
