Page 483 - The Toxicology of Fishes

The Endocrine System 463

Actions of Hormones at Target Tissues
Activation of Intracellular Second Messengers
Binding of peptide, protein, and steroid hormones to cell-surface receptors on target cells activates
intracellular signal transduction systems, leading to the hormonal response. Binding of GnRH to a
specific GPCR on gonadotropes activates several signal transduction pathways in the three teleost models
investigated in detail: goldfish, salmonids, and tilapia (Ando et al., 2001; Chang and Jobin, 1994; Yaron
et al., 2001). GnRH causes rapid activation of a G-protein (G q/11 ) and a second-messenger cascade
involving phospholipase C (PLC) and the formation of inositol triphosphate (IP ) and diacylglycerol
3
(DAG). DAG causes activation of protein kinase C (PKC), whereas IP has been shown to cause
3
intracellular calcium concentrations to increase in mammalian gonadotropes due to mobilization from
internal stores. GnRH also induces calcium influx through voltage-sensitive calcium channels (VSCCs)
into fish gonadotropes. The increase in intracellular calcium concentrations seems to be of primary
importance in the stimulation of gonadotropin secretion, whereas both calcium levels and activation
PKC are involved in GnRH regulation of gonadotropin subunit gene expression (Ando et al., 2001; Gur
et al., 2001). Moreover, recent evidence suggests that PKC acts via stimulation of mitogen-activated
protein kinase (MAPkinase). In addition, phospholipase A and arachidonic acid appear to be involved
2
in gonadotropin release from tilapia pituitaries. Also, a cAMP-dependent protein, protein kinase A (PKA),
is activated by GnRH in tilapia (Yaron et al., 2001). The signal transduction systems activated by
gonadotropins are highly conserved among the vertebrates. The adenylate cyclase system is of central
importance in mediating the steroidogenic actions of gonadotropins in fish as well as in other vertebrates
(Van Der Kraak and Wade, 1994). Cyclic AMP levels increase markedly in spotted seatrout ovaries
within 15 minutes of gonadotropin stimulation in vitro, prior to the increase in steroid synthesis (Singh
and Thomas, 1993). Calcium from both intracellular and extracellular pools is also an important com-
ponent of the signaling pathway (Benninghoff and Thomas, 2005). The calcium-binding protein calm-
odulin participates in the regulation of intracellular calcium levels, whereas VSCCs regulate calcium
influx. Other second messengers such as PKC, inositol-1,4,5-triphosphate, diacylglycerol, and arachi-
donic acid appear to be components of the gonadotropin signaling pathway in certain steroidogenic
tissues (Van der Kraak and Wade, 1994). Therefore, chemicals could potentially interfere with a variety
of signal transduction systems to modulate the actions of gonadotropin.

Genomic Steroid Actions via Nuclear Steroid Receptors
The classic genomic mechanism of steroid action involves diffusion of steroids into the cell where they
bind to intracellular receptors (nuclear steroid receptors), resulting in receptor activation and binding to
hormone response elements on genes (in conjunction with other transactivating proteins) to alter their
rates of transcription (Figure 10.2) (Beato and Sanchez-Pacheco, 1996; Tsai and O’Malley, 1994). The
nuclear steroid receptors—estrogen (ER), androgen (AR), progestogen (PR), glucocorticoid (GR), and
mineralocorticoid (MR)—are members of the nuclear receptor superfamily of ligand-activated transcrip-
tion factors that includes receptors for thyroid hormone, vitamin D, ecdysone, and retinoids as well as
many orphan receptors for which the ligands are currently unknown (Laudet, 1997). Nuclear steroid
receptor proteins have a similar modular structure with six functional domains (A to F). At the N-terminal
end, the hypervariable A/B domains possess sequences required for transcriptional activation. The DNA-
binding region (C domain) is the most highly conserved region of the receptor protein (60 to 95%
homology) and has two protruding zinc fingers that enable the ligand-activated receptor to bind to
hormone response elements (specific DNA sequences in the regulatory regions of genes) of the target
genes. The variable hinge region (D domain) is involved in nuclear localization. The large ligand-binding
domain (LBD), or E domain, is conserved and displays a high degree of sequence homology, particularly
with members of the same receptor subfamily (38 to 60% sequence identity between teleostean and
mammalian ERs) (Hawkins et al., 2000). In addition, the LBD is involved in receptor dimerization, heat
shock protein binding, and transcriptional activation. At the C-terminal end, the F domain is highly
variable and of unknown function.