T4 in serum, Decreased leads to Reduced, Anterior swim bladder inflation

Reduced anterior chamber inflation upon disruption of the thyroid hormone system is in most cases, but not always, accompanied by reduced whole body T3 levels. Stinckens et al. (2016) found a consistent relationship between reduced whole body T4 levels, but not T3 levels, and reduced anterior chamber inflation after exposure to 2-mercaptobenzothiazole (MBT). Possibly, local T4 levels in the swim bladder tissue were too low to allow for enough local activation to T3. This relates to the general uncertainty on serum versus tissue TH levels. Alternatively, differences in timing between T3/T4 measurements (at 120hpf and 32dpf), the moment when there is a need for T3 to inflate the swim bladder (unknown but probably in between 120hpf and 32dpf) and the observation of the phenotype (32dpf), could lead to the hypothesis that T3 concentration was reduced in between the two measurements. There is also a possibility that the effect of MBT on anterior chamber inflation is not directly caused by decreased thyroid hormone levels, but rather by another mechanism such as oxidative stress. MBT is known to elevate the production of reactive oxygen species (ROS) levels in fish cells (Zeng et al., 2016). In general, chemicals may have multiple modes of action and effects on autophagy, ROS, cardiac function may impact swim bladder inflation.

The mechanism through which reduced T4 hormone concentrations in serum result in anterior chamber inflation impairment is not yet understood. The anterior chamber is formed by evagination from the cranial end of the posterior chamber (Robertson et al., 2007, Winata et al., 2009). Several hypotheses could explain effects on anterior chamber inflation due to reduced T4 levels:

• Evagination from the posterior chamber could be impaired. Villeneuve et al. (unpublished results) showed that although the anterior bud was present after exposure to a deiodinase 2 inhibitor, the anterior chamber did not inflate.
• The formation of the tissue layers of the anterior swim bladder could be affected, although Villeneuve et al. (unpublished results) observed intact tissue layers of the anterior swim bladder after exposure to a deiodinase 2 inhibitor.
• The anterior chamber is inflated with gas from the posterior chamber through the communicating duct. Impaired gas exchange between the two chambers could be at the basis of impaired anterior inflation. Both Nelson et al. (2016) and Stinckens et al. (2016) found that posterior chambers were larger when anterior chambers were smaller or not inflated at all. The sum of the areas of the posterior and anterior chambers remained constant independent of inflation of the anterior chamber (Stinkens et al., 2016). These results suggest retention of the gas in the posterior chamber.
• Since gas exchange relies on a functional communicating duct between the posterior and anterior chamber, and the communicating duct is known to progressively narrow and eventually close during development, a dysfunctional communicating duct or a closure prior to anterior inflation could inhibit inflation. However, Villeneuve et al. (unpublished results) showed that the communicating duct was anatomically intact and open after exposure to iopanoic acid (a deiodinase 2 inhibitor), still leading to impaired anterior inflation.
• Lactic acid production which is essential for producing gas to fill the swim bladder could be affected, although the observation that the total amount of gas in both chambers is not affected when anterior inflation is impaired seems to contradict this (Stinckens et al., 2016).
• Possibly there is an effect on the production of surfactant, which is crucial to maintain the surface tension necessary for swim bladder inflation.
• Reinwald et al. (2021) showed that T3 and propylthiouracil treatment of zebrafish embryos altered expression of genes involved in muscle contraction and functioning in an opposing fashion. The authors suggested impaired muscle function as an additional key event between decreased T3 levels and reduced swim bladder inflation.

In some cases indirect effects may play a role in the impact of chemical exposure or genetic knockdown/knockout on swim bladder inflation. For example, dual oxidase also plays a role in oxidative stress.

Reduced anterior chamber inflation upon disruption of the thyroid hormone system is consistently accompanied by reduced whole body T4 levels when fish are exposed to thyroid hormone synthesis inhibitors. however, when fish are exposed to deiodinase inhibitors, in the absence of feedback processes, stable T4 levels would be expected. Stable T4 levels were indeed observed in 14, 21 and 32 day old zebrafish exposed to iopanoic acid, a deiodinase inhibitor. While Cavallin et al. (2017) found a consistent relationship between reduced whole body T3 levels, and reduced anterior chamber inflation after exposure of fathead minnows to iopanoic acid, they observed increased T4 levels. Possibly, the inhibition of the conversion of T4 to T3 resulted in a compensatory mechanism that increased T4 levels. This was accompanied by increases of deiodinase 2 and 3 mRNA levels, which also indicate a compensatory response to deiodinase inhibition.