Chapter 1 – General Introduction
1.4 Photoperiodism reinvented: TH and DIO expression
In zebrafish, pineal melatonin is used as a systemic signal for circadian and seasonal light/dark cycles (Cahill, 1996). Melatonin is circulated through the cerebrospinal fluid and blood, acting on targets throughout the body, through tissue‐specific expression on melatonin receptors (Vatine et al., 2011). Specific photoperiodic effects on growth and reproduction may be determined by
melatonin modulation of control centres such as the pituitary, hypothalamus and the gonads themselves (Hazlerigg & Wagner, 2006). The photoperiodic expression of melatonin receptors in a range of target tissues has been explored in detail in the current work and will be discussed in detail in chapter 2.
1.4 Photoperiodism reinvented: TH and DIO expression Increasingly evidence suggests that thyroid hormone (TH) is crucial for the
expression of seasonal rhythms in vertebrates (Barrett et al., 2007). TH expression is controlled by hypothalamic neurosecretory cells releasing thyroid‐stimulating hormone‐releasing hormone (TSH‐RH). This releasing hormone targets the thyrotroph cells of the anterior pituitary, increasing the release of thyroid‐
stimulating hormone (TSH), which stimulates the production of thyroxine (T4) from the thyroid gland (Nakao et al., 2008b).
Vertebrate thyroid hormones have two isoforms; thyroxine (T4) is circulated systemically, with a low biological activity; and the more biologically potent form, triiodothyronine (T3), is created in target tissues through the local conversion of T4 by deiodinase enzymes (Hazlerigg & Wagner, 2006). As shown in figure 1.8, the two principal deiodinase enzymes found in local tissues are type 2 deiodinase (Dio2) and type 3 deiodinase (Dio3) (Hazlerigg & Wagner, 2006). Dio2 catalyzes
the conversion of T4 to the active T3 form, while Dio3 converts T3 to inactive reverse‐T3 (rT3) form. The relative expression and activity of Dio2 and Dio3 determine the levels of biologically active T3 in the brain (Nakao et al., 2008b).
Figure 1.8 ‐ Thyroid hormone (TH) processing by local deiodinase enzymes. The main form of circulating thyroid hormone, T4 is converted to T3 in tissues expressing type 2 deiodinase (Dio2). T4 can also be
converted to an inactive form, reverse T3 (rT3) by type 3 deiodinase (Dio3). Both rT3 and T3 can be processed into T2. Figure from Hazlerigg and Loudon, 2008.
Using autumn‐breeding mammals such as sheep, Fred Karsh and colleagues have shown that thyroidectomy removes the seasonal (or photo‐refractory) inhibition of reproductive behavior normally expressed in spring (Billings et al., 2002).
Injections of T4 in these thyroidectomised animals restores endogenous springtime reproductive inhibition, but has only a weak effect on the onset of photoperiodic reproduction in autumn (Billings et al., 2002). These inhibitory springtime T4 effects have been shown to be maximally effective within the basal hypothalamus of thyroidectomised ewes (Anderson et al., 2003). Experiments using microinjection mapping have shown the ependymal cells of the ventral hypothalamus as the site of the greatest photoperiodic induced Dio2/Dio3 activity in both long and short day breeding mammals (Hanon et al., 2008). This
ependymal cell layer is composed of tanycyte cells surrounding the 3rd ventricle which act as regulatory cells for the transport of solutes in/out of the brain (Hazlerigg & Loudon, 2008). These tanycyte cells project to the pars tuberalis of the anterior pituitary, and may regulate the hypothalamic‐pituitary hormone relay system (Nakao et al., 2008b).
Figure 1.9 gives a schematic drawing illustrating the differences between mammalian and non‐mammalian (vertebrate) light reception systems, and a model of the connection between the hypothalamus and pituitary gland specifically.
Figure 1.9: Schematic drawing
highlighting the proposed differences between vertebrate/avian and
mammalian photo‐neuroendocrine systems.
A) Coronal section; (left) in ancestral systems, light input (red) to different structures serves different functions such as vision, circadian and photoperiodic input. These signals are integrated and initiate photoperiodic‐linked endocrine secretion from the pituitary (open
arrows). In mammals (right), light input is through the eyes, and the hypothalamus integrates these cues into photoperiodic information. These signals may be translated in the pars tuberalis, before passing to the pituitary gland where endocrine output is initiated.
B) Schematic of the hypothalamic‐pituitary portal system. Hypothalamic
neurosecretory cells release signals into median eminence (ME) capillaries, which drain into portal vessels (red) to the pituitary endocrine cells, whose hormones are secreted into peripheral circulation. The dashed line indicates the plane of section of the inset, a coronal view of the ME ventral to the third ventricle (3V); cells surrounding the 3V form the paraventricular zone (PVZ), extending to the ME. The
Working with Japanese quail (C. japonica), Brian Follett and others have shown that thyroidectomy inhibits seasonal responses to photoperiod, such as gonad growth, and this response can be restored by T4 injections (Follett & Nicholls, 1985). T4 levels in the hypothalamus increase during the lengthening days of spring, and drop in autumn, reflecting the stimulating long day photoperiod of the species (Yoshimura et al., 2003). When the quail were exposed to stimulatory long photoperiods, Dio2 was significantly up regulated in the hypothalamus,
synthesizing more active T3 (Yasuo et al., 2005). Exposure to inhibitory short photoperiods suppressed Dio2 while increasing Dio3 expression, effectively limiting T3 availability in a seasonal manner (Nakao et al., 2008b). T3 targets include the GnRH cells of the median eminence (in the ventral hypothalamus), which release GnRH pulses into the portal blood supply, managing the subsequent release of downstream pituitary hormones such as LH and FSH (Nakao et al., 2008b).
Experiments in mammalian models with both long and short day breeders suggest that T4/T3 expression by local Dio enzymes conversion governs seasonal
reproduction (Hazlerigg & Loudon, 2008) and the seasonal photoperiod regulates the expression of these deiodinase enzymes within the basal hypothalamus (Nakao et al., 2008b).
Figure 1.10: Schematic drawing of the proposed photoperiodic regulation of thyroid hormones in avian ependymal cells. Light received by deep brain
photoreceptors induces TSH expression in the pituitary pars tuberalis (PT; pink).
Long‐day expression of TSH leads to the formation of TSH in the PT, acting on TSH receptors (TSHR) expressed by ependymal cells (blue); leading to seasonal
changes in hypothalamic Dio1‐2‐3 expression. Tanycytes convert T4 into bio‐active T3, stimulating the release of hypothalamic GnRH. GnRH is carried to the pituitary (pars distalis; PD) by portal blood vessels, modulating release of LH and FSH from the anterior pituitary into general circulation. Figure adapted from Nakao et al., 2008.
The principal components making up the thyroid axis are conserved across vertebrates, including teleost fish (Power et al., 2001). As with mammalian and avian models, thyroid hormones play crucial roles in regulating development, differentiation and metabolism and species are unable to grow and mature normally without them (Porterfield & Hendrich, 1993). Much of the past TH fish‐
specific literature has highlighted the importance of THs during metamorphosis, early development and growth (Power et al., 2001). Thyroid hormone receptors (TR) have been isolated from several teleosts such as Japanese flounder
(Paralichthys oliaceus), sea bream (Sparus aurata) and zebrafish (D. rerio) (Power
In Atlantic cod (G. morhua) photoperiod alterations affect cyclic patterns of sex steroids, thyroid hormones, and the timing of spawning (Norberg et al., 2004). In this species, plasma T3 levels are highest from mid‐summer to fall, coinciding with the highest annual growth rate (Norberg et al., 2004). Both plasma T3 and T4 levels exhibit a strong seasonal rhythm in teleosts and are associated with the uptake of thyroid hormones by developing eggs from circulating plasma levels, accounting for significant declines in plasma T3 seen prior to spawning (Tagawa & Brown, 2001). Previous data suggested temperature acted as a major regulator of TH expression, but modern studies have focused on photoperiod as a primary
entrainer of thyroid hormone secretion (Comeau et al., 2000). Thyroid hormones also act as growth inducers in fish (Donaldson et al., 1979), stimulating growth hormone secretion (Ebbesson et al., 1998). Up till now there has been no literature available regarding the seasonal photoperiodic T4/T3 expression by local Dio enzymes within the basal hypothalamus in light entrained zebrafish and the work presented here (Chapter 3; figures 3.1 and 3.3) is the first of its kind.