322
Marine Mammal Physiology: Requisites for Ocean Living
the kidney and the liver of rats has been observed for several decades (Wiśniewska et al. 1970;
Trojanowska and Sapota 1974), and species of Hg have been clearly demonstrated to induce
MTs in other mammalian models. Indeed, it was assumed that sequestration by MT is a main
detoxification function for mercury (Roesijadi 1992). However, studies in marine mammal
species have indicated that a relatively small percentage of mercury is bound to MTs in the
liver and kidneys. For instance, one study found that the percentage of tissue Hg bound to
metallothioneins ranged from 14.5% to 20.0% in the kidney and 2.0% to 3.2% in the liver of
California sea lions (Zalophus californianus) (Lee et al. 1977). Similarly, low percentages of Hg
bound to MT have been found in narwhals (5% in the liver, 10% in the kidney, Wagemann
et al. 1984), and pilot whales (79% of THg in the insoluble fraction, Caurant et al. 1996). It
is possible that MTs represent an acute response to intracellular MeHg + or Hg 2+ , and that
transient sequestration via binding to MTs facilitates transport to other organs or eventual
biotransformation of metals into inactive forms (e.g., tiemmanite).
The low percentages of Hg bound to MTs are intriguing because it has been proposed that
toxic effects of heavy metals occur when free metals exceed the binding capacity of MTs, or
other high-affinity scavenging proteins. The spillover hypothesis was first proposed by Winge
et al. (1974) who showed that pathologic effects of Cd were only seen when free Cd entered
other protein fractions. The situation becomes more complex, however, when considering
that there is more than one metal present and these metals have different concentrations and
binding affinities for the numerous isoforms of MT proteins (Hamilton and Mehrle 1986).
Interestingly, the expression of MT mRNA has been shown to increase following exposure to mercury chloride (Hg 2+ ), accompanied by an increase in the MT concentration (Reus
et al. 2003). The authors speculate that MTs bind Hg due to the high concentration of sulfhydryl groups (–SH) on cysteine residues, and that the MT–Hg complex enters the nucleus
where it disassociates and attaches the metal response element of the MT gene, resulting
in an increase in the MT expression. The mammalian MT gene has several 5′ upstream elements that allow it to responds to stimuli, one of them being the metal response element
(MRE). The MREs, in the presence of heavy metals such as Zn and Cd, respond to metalregulatory transcription factors such as MT1-F, which has been described in a number of
vertebrate species (Andrews 2000). Another explanation for the induction of MTs is the role
they play in mitigating cellular oxidative stress. Metallothionein mRNA expression has
been shown to be induced not only by heavy metals, but by strongly oxidizing agents such
as hydrogen peroxide (Andrews 2000). Mercury is known to cause oxidative stress, thus the
transcriptional response of cells to Hg might be occurring in response to oxidative damage
rather than Hg bound to MT, or it could be a combination of the two mechanisms.
Glutathione peroxidase is an endogenous antioxidant which prevents both lipid peroxidation as well as catalyzes hydrogen peroxide into water. Other antioxidant enzymes
(and cofactors) have been shown to rescue rats from lipid peroxidation following chronic
Cd exposure (in this case N-acetylcysteine or Vitamin E were co-delivered with Cd),
resulting in protection from renal toxicity (Shaikh et al. 1999). Diving mammals have
been shown to have higher activities of GPx compared to terrestrial mammals (Wilhelm
Filho et al. 2002). Similar results were demonstrated with higher GPx activity in ringed
seals versus domestic pig in the heart, the lung, and the muscle tissue (Vázquez-Medina
et al. 2006). The higher activity of antioxidant defenses as an adaptation to diving physiology has been discussed earlier in this chapter, however, there is no reason not to suspect
that higher activities of the enzymes could not provide, at least in part, protective effects
against heavy metal-induced oxidative stress as well. In fact, it is more difficult to speculate that increased GPx activity would be independent of some role in poison detoxification or biotransformation.
Marine Mammal Physiology: Requisites for Ocean Living
the kidney and the liver of rats has been observed for several decades (Wiśniewska et al. 1970;
Trojanowska and Sapota 1974), and species of Hg have been clearly demonstrated to induce
MTs in other mammalian models. Indeed, it was assumed that sequestration by MT is a main
detoxification function for mercury (Roesijadi 1992). However, studies in marine mammal
species have indicated that a relatively small percentage of mercury is bound to MTs in the
liver and kidneys. For instance, one study found that the percentage of tissue Hg bound to
metallothioneins ranged from 14.5% to 20.0% in the kidney and 2.0% to 3.2% in the liver of
California sea lions (Zalophus californianus) (Lee et al. 1977). Similarly, low percentages of Hg
bound to MT have been found in narwhals (5% in the liver, 10% in the kidney, Wagemann
et al. 1984), and pilot whales (79% of THg in the insoluble fraction, Caurant et al. 1996). It
is possible that MTs represent an acute response to intracellular MeHg + or Hg 2+ , and that
transient sequestration via binding to MTs facilitates transport to other organs or eventual
biotransformation of metals into inactive forms (e.g., tiemmanite).
The low percentages of Hg bound to MTs are intriguing because it has been proposed that
toxic effects of heavy metals occur when free metals exceed the binding capacity of MTs, or
other high-affinity scavenging proteins. The spillover hypothesis was first proposed by Winge
et al. (1974) who showed that pathologic effects of Cd were only seen when free Cd entered
other protein fractions. The situation becomes more complex, however, when considering
that there is more than one metal present and these metals have different concentrations and
binding affinities for the numerous isoforms of MT proteins (Hamilton and Mehrle 1986).
Interestingly, the expression of MT mRNA has been shown to increase following exposure to mercury chloride (Hg 2+ ), accompanied by an increase in the MT concentration (Reus
et al. 2003). The authors speculate that MTs bind Hg due to the high concentration of sulfhydryl groups (–SH) on cysteine residues, and that the MT–Hg complex enters the nucleus
where it disassociates and attaches the metal response element of the MT gene, resulting
in an increase in the MT expression. The mammalian MT gene has several 5′ upstream elements that allow it to responds to stimuli, one of them being the metal response element
(MRE). The MREs, in the presence of heavy metals such as Zn and Cd, respond to metalregulatory transcription factors such as MT1-F, which has been described in a number of
vertebrate species (Andrews 2000). Another explanation for the induction of MTs is the role
they play in mitigating cellular oxidative stress. Metallothionein mRNA expression has
been shown to be induced not only by heavy metals, but by strongly oxidizing agents such
as hydrogen peroxide (Andrews 2000). Mercury is known to cause oxidative stress, thus the
transcriptional response of cells to Hg might be occurring in response to oxidative damage
rather than Hg bound to MT, or it could be a combination of the two mechanisms.
Glutathione peroxidase is an endogenous antioxidant which prevents both lipid peroxidation as well as catalyzes hydrogen peroxide into water. Other antioxidant enzymes
(and cofactors) have been shown to rescue rats from lipid peroxidation following chronic
Cd exposure (in this case N-acetylcysteine or Vitamin E were co-delivered with Cd),
resulting in protection from renal toxicity (Shaikh et al. 1999). Diving mammals have
been shown to have higher activities of GPx compared to terrestrial mammals (Wilhelm
Filho et al. 2002). Similar results were demonstrated with higher GPx activity in ringed
seals versus domestic pig in the heart, the lung, and the muscle tissue (Vázquez-Medina
et al. 2006). The higher activity of antioxidant defenses as an adaptation to diving physiology has been discussed earlier in this chapter, however, there is no reason not to suspect
that higher activities of the enzymes could not provide, at least in part, protective effects
against heavy metal-induced oxidative stress as well. In fact, it is more difficult to speculate that increased GPx activity would be independent of some role in poison detoxification or biotransformation.