Level 1-9 ( Figure 1 ) demonstrates a summary of contributions of Dratman regarding development of the concepts of nongenomic action of TH in adult mammalian brain. These studies are summarized below ( Figure 2 ). She provided insights into TH actions including (a) the adult brain responsiveness to TH, nullifying the earlier concept of non-responsiveness of adult brain to TH, (b) the maintenance of adult brain TH levels by deiodinase systems, and (c) iodothyronine production as catecholamine analogs and their possible action in brain, particularly in the case of thyroidal illness (16). Based on the experimental evidence since the 1970s ( Figure 1 ), Dratman proposed the concept of nongenomic action of TH in adult brain [i.e., the sympathomimetic activity of “iodothyronine-derived neurohormones” in brain (17). Dratman identified, for the first time, the in vivo accumulation of radiolabeled T4/T3 at nerve terminals using synaptosomes isolated from adult rat brains after intravenous injections of T3/T4 ( Figure 2 ) (6, 18–20). Additionally, her group reported that T4/T3 might act peripherally in the salivary gland (21) and centrally in autonomic nervous system for regulation of blood pressure (20) and heart rate (22).

Behavioral studies showed that TH might influence the circadian rhythm of temperature regulation (23) and have an impact on affective disorders (24) related to thyroidal dysfunctions.

Dratman extended her studies with findings of the in vivo localization of radiolabeled T4/T3 in different regions of adult rat brain (a) after injection of TH (6, 9, 19, 20, 25–31), (b) during altered thyroidal states (20, 32–37) and (c) after pharmacologically altered adrenergic systems in brain during euthyroid (6, 28, 34–36) and altered thyroid conditions (6, 34–37). The pharmacological interventions in their studies were induced by adrenergic uptake inhibitors such as desmethylimipramine (35–37) and reserpine (28), and the neurotoxic agent [N-(2-chloroethyl)-N-2-bromobenzylamine hydrochloride (DSP-4)] selective for adrenergic neurons in the locus coeruleus (6, 34). Notably, this series of reports supported the concept of accumulation of T3 in different regions of adult mammalian brain and its action as neurotransmitter and/or co-transmitter with adrenergic systems ( Figure 2 ).

Dratman further examined brain homeostatic mechanisms to maintain the T3/T4 levels at nerve terminals under altered conditions (9, 33, 35, 37). Finally, she proposed the possibilities of postsynaptic action of THs or derivatives after their release from adrenergic nerve terminals (34, 38). In addition, her group reported that T3 can induce nongenomic action on neuronal activation of hippocampal cells, one of the target regions of adrenergic system in adult rat brain (14). Such findings help to explain the physiological or pathophysiological influences of TH in psychobehavioral control in adult mammalian brain.

Separate in vivo and in vitro studies have been executed by other investigators using synaptosomes isolated from adult rodent brains, to find the exact mechanism of nongenomic action(s) of T3 at nerve terminals (see Figure 1 : levels 10-19, Figure 2 ). These studies described several nongenomic actions exerted by TH, including protein phosphorylation, calcium-flux, NOS activity, putative membrane receptor binding, uptake and release of THs. Synaptosomes isolated from the whole brains of adult rats show deiodinase activities for conversion of T4 to rT3 and T3 to 3,3’T2 or 3’,5’T2 (39). T4 and T3 have been estimated in synaptosomes isolated from 11 different regions of adult rat brain (40). The adult rat brain shows thyroid homeostatic mechanisms at nerve terminals by increasing synaptosomal T4 and T3 levels (41, 42) during altered thyroidal conditions, particularly at initial stages of the altered conditions (43). Anti-depressant treatments alter T3 levels in synaptosomes isolated from frontal cortex (44), amygdala (45) and cortical areas (46) of adult rat brains. Therefore, these in vivo studies using isolated synaptosomes under different experimental conditions indicate that nerve terminals of adult mammalian brain have a capacity to maintain their T4/T3 levels for a certain extent of conditions.

In adult rat brain synaptosomal fractions, a Na⁺-dependent carrier-mediated uptake for T3 was demonstrated that involved both high-affinity and low-affinity transport systems. T4 was transported by a concentration-dependent but Na⁺-independent manner (47). One study showed that cortical synaptosomes can release T3, but not T4, under depolarized conditions through a Ca²⁺-dependent process, thus supporting the concept that T3 can act as a neurotransmitter (48). Still, additional research is needed to confirm this point. Furthermore, T3 enhances Ca²⁺-dependent release of GABA under depolarizing conditions (49). T3 enhances Na⁺-dependent tryptophan transport (50) and inhibits leucine (51, 52) or GABA (53) uptakes in synaptosomes. These in vitro studies show that T3 is transported at nerve terminals and/or alters the ion-dependent transport of amino acids and amino acid derivatives as rapid nongenomic actions of T3 at nerve terminals of adult mammalian brain.

T3 shows high-affinity binding to synaptosomal membranes (54–56) isolated from adult rat brain cerebral cortex. T3 inhibits Na⁺/K⁺-ATPase activity in cerebrocortical synaptosomal membranes (56) and thus can modulate the neuronal depolarization in adult rat brain (57, 58). T3 inhibits membrane bound ectonucleotidase in synaptosomes isolated from hippocampus of adult rat brain and modulates ATP hydrolyses (59). These studies indicate that T3 can act in a nongenomic fashion to minimize the ATP loss at the synaptic level. T3 stimulates the activities of Ca²⁺/Mg²⁺-ATPase (60) and acetylcholinesterase (61) in cerebrocortical synaptosomes. Hence, T3 may have a role in calcium homeostasis and the modulation of cholinergic neurotransmission in adult brain. T3 inhibits glutamate-induced Ca²⁺-uptake in synaptosomes isolated from mouse whole brain (62). Furthermore, T3 enhances depolarization-induced Ca²⁺-uptake (63) in synaptosomes isolated from rat cerebral cortex and causes a transient rise in intrasynaptosomal Ca²⁺-calcium levels (64) which indicates the nongenomic action of T3 on the Ca²⁺-dependent neurotransmission process. Altered thyroidal conditions also mobilize the synaptosomal Ca²⁺-level (65). Interestingly, the transient rise of intrasynaptosomal Ca²⁺-was found to be associated with synaptosomal nitric oxide synthase activation (64). This indicates that T3 can act through a nongenomic Ca²⁺-calcium-dependent nitric oxide (NO) signaling pathway at synaptic regions and thereby modulate neurotransmission. In addition, a series of in vitro experiments demonstrated the T3-induced Ca²⁺- and calmodulin-dependent synaptosomal protein phosphorylation (66–68) that underlies nongenomic cellular signaling pathway ( Figure 2 ).

As mentioned in section 3.1.1, much of Dratman’s research investigated the anatomical and subcellular localization of radioactivity in adult rodent brain following injections of labeled THs (8,9,19, 20,25-31). Electron microscopic studies showed accumulation of radiolabeled THs in neuropil, especially in nerve terminal regions (9, 20, 28). In addition, subcellular fractionation by differential centrifugation of homogenates of brain showed that the radiolabeled THs were concentrated in synaptosomes (6, 18–20).

The subcellular localization of THs points to the potential role of the compounds at the synapse. Dratman therefore espoused the hypothesis that THs (or their derivatives) might have neurotransmitter-like actions. The localization in nerve terminals readies the compounds for release into the synaptic cleft, where they might have influences on postsynaptic or presynaptic receptors. Thus, the THs are optimally positioned to participate in a synaptic signaling role.

Thaw-mount autoradiography indicated that following intravenous (IV) administration of ¹²⁵I-T3, radioactivity corresponding to T3 (80%) or other iodinated organic compounds (15%) accumulated in discrete brain regions. At 10 hours post-injection, the radiolabel shifted to fiber tracts, implying that it is transported along axons (25). Additional studies showed that IP administration of the DSP-4, toxin specific to adrenergic neurons in locus coeruleus (LC), reduced the distribution of T3 immunohistochemistry in specific sites in the forebrain, the target site of the adrenergic neurons originated from LC (34). These data suggest that DSP-4 disrupted the transport of T3 throughout the brain and indicate that the transport is orthograde. Since orthograde axonal transport requires energy (69), such transport of T3 may indicate the importance of the hormone for actions at the nerve terminal, in keeping with a signaling function at the synapse.

The concentration of radioactivity in discrete brain regions following administration of radiolabeled THs suggests that the actions of the hormones or their derivatives are specific to particular brain functions under particular circumstances in adult mammalian brain. These actions could be either nongenomic or genomic.