Fyziologie působení farmak a
toxických látek
Přednáška č.7
Principy endokrinní regulace a
endokrinní disrupce u
bezobratlých
HormonHormonáálnlníí regulace u bezobratlých:regulace u bezobratlých:
Hormony uHormony u ččlenovclenovcůů::
Hormony u ostnokoHormony u ostnokožžccůů a pla plášášttěěncncůů::
Modes of insect development. Molts
are represented as arrows.
(A) Ametabolous (direct)
development in a silverfish. After a
brief pronymph stage, the insect
looks like a small adult.
(B) Hemimetabolous (gradual)
metamorphosis in a cockroach.
After a very brief pronymph phase,
the insect becomes a nymph. After
each molt, the next nymphal instar
looks more like an adult, gradually
growing wings and genital organs.
(C) Holometabolous (complete)
metamorphosis in a moth. After
hatching as a larva, the insect
undergoes successive larval molts
until a metamorphic molt causes it
to enter the pupal stage. Then an
imaginal molt turns it into an adult.
MetamorfMetamorfóóza hmyzu:za hmyzu:
Regulation of insect
metamorphosis. (A)
Structures of juvenil
hormone, ecdysone,
and the active moltin
hormone 20hydroxyecdysone.
(B
General pathway of
insect metamorphosis
Ecdysone and juvenile
hormone together
cause molts to keep
the status quo and
form another larval
instar. When there i
a lower concentration
of juvenile hormone,
the ecdysone-induced
molt produces a pupa
When ecdysone acts
in the absence of
juvenile hormone, the
imaginal discs
differentiate, and
the molt gives rise t
the adult
Another candidate for the
JH receptor role is the
Methoprene-tolerant (Met)
Per-Arnt-Sim (PAS)
domain protein, whose loss
confers tolerance to JH and
its mimic methoprene in the
fruit fly Drosophila
melanogaster. (Konopová et
Formation of the ecdysone receptors. Alternative mRNA splicing of the
ecdysone receptor (EcR) transcript creates three types of EcR mRNAs. These
generate proteins having the same DNA-binding site (blue) and
hydroxyecdysone-binding site (red), but with very different amino termini.
Ec receptory:Ec receptory:
Three isoforms of EcR have been identified in insects, each with a different,
stage-specific role in regulation of molting and development. This allows for one
steroid hormone to induce a variety of different tissue responses. In general,
EcR A is predominant when cells are undergoing a maturation response (from
juvenile to adult) and is predominant in imaginal discs, whereas EcR B1
predominates in juvenile cells during proliferation or regression. Little is known
about the function of the EcR B2 isoform.
DNA and hormone binding are similar in the three isoforms of EcR. Little is
known about the crustacean EcR isoforms and how they change during the molt
cycle. However, the EcR that has been cloned from the crab, Uca pugilator
(U31817, GenBank), shares 85­87% homology with that of Drosophila (M74078,
GenBank). The differences are primarily in the region of the molecule involved
with dimerization. Similar sequence similarities are found between the
heterodimeric partner, USP.
There are several ecdysteroids which bind EcR, including 20-hydroxyecdysone,
turkesterone, makisterone A, ponasterone A, and muristerone A. Some
arthropods may use specific ecdysteroids as their principal molting hormone, but
often several ecdysteroids are found within one group. The primary molting
hormone for a range of organisms, including some insects and crustacea, is 20OH
ecdysone (20 HE). Among other examples, makisterone A is an important
hormone for some crustacea and hemipteran insects.
Hydroxyecdysone-induced puffs in cultured
salivary gland cells of D. melanogaster. (A)
Uninduced control. (B-E) Hydroxyecdysonestimulated
chromosomes at (B) 25 minutes,
(C) 1 hour, (D) 2 hours, and (E) 4 hours.
The Ashburner model of hydroxyecdysone
regulation of transcription.
Hydroxyecdysone binds to its receptor,
and this compound binds to an early puff
gene and a late puff gene. The early puff
gene is activated, and its protein product
(1) represses the transcription of its own
gene and (2) activates the late puff gene,
perhaps by displacing the ecdysone
receptor. (After Richards 1992.)
Struktury Ec hormonStruktury Ec hormonůů::
Cholesterol (from diet- a vitamin for insects)
Prothoracic
gland
20-HydroxyecdysoneEcdysone
mono-oxygenase
(fat body, epidermis)
Conjugates
(storage)
Metabolites
(excretion)
Synthesis of molting hormonesSynthesis of molting hormones
Prothoracicotropic hormone (PTTH)
Protein of 30-kDa active as a homodimer linked by
a disulfide bond
Produced by two pairs of lateral neurosecretory
cells
Released from corpora
allata (moths) or
corpora cardiaca
(most insects)
PTTH cells of Manduca visualized
with anti-PTTH antibody
Ecdysone synthesis and secretion are initiated by prothoracicotropic
hormone (PTTH), a hormone produced by two pairs of neurosecretory
cells in the brain. PTTH was first isolated and characterized from the
silkmoth Bombyx mori. PTTH has conserved seven cysteine residues,
several hydrophobic regions and an N-glycosylation site. Only the
homodimeric form of Bombyx PTTH is biologically active. Bombyx
PTTH is thought to be a member of the transforming growth factor(TGF-)
family.
The role of PTTH
Initiates every molt
­ stimulates prothoracic glands to synthesize and release
ecdysone
­ serves as "mission control" allowing molt if the conditions
are right
* factors affecting decision to molt are species-specific
­ stretch of the abdomen by a blood meal in Rhodnius
­ completion of the cocoon in some moths
­ escape from wet diet in flies
Phytoecdysteroids (PEs) are a family of about 200 plant steroids
related in structure to the invertebrate steroid hormone 20hydroxyecdysone.
Typically, they are C27, C28 or C29 compounds
possessing a 14a-hydroxy-7-en-6-one chromophore and A/B-cis ring
fusion (5-H).
PEs are attracting renewed attention because of their specific
effects on invertebrate development (potential in invertebrate pest
control) and their varied benign pharmacological actions on mammals
(biomedical applications and gene switches). In the past three
decades, several thousand species of plants have been surveyed for
the presence of PEs and the structures of over 200 PEs have been
deduced. The most frequently encountered PE is 20E, the principal
physiological inducer of moulting and metamorphosis in arthropods.
Fytoekdosteroidy:Fytoekdosteroidy:
SyntSyntééza ekdysteroidza ekdysteroidůů::
SyntSyntééza ekdysteroidza ekdysteroidůů aa úúloha cytochromloha cytochromůů
P450:P450:
SyntSyntééza ekdysteroidza ekdysteroidůů aa úúloha cytochromloha cytochromůů P450:P450:
JuvenilnJuvenilníí hormon a jeho analogy:hormon a jeho analogy:
Regulation of insect
metamorphosis. (A)
Structures of
juvenile hormone,
ecdysone, and the
active molting
hormone 20hydroxyecdysone.
(B
General pathway of
insect
metamorphosis.
Ecdysone and
juvenile hormone
together cause molt
to keep the status
quo and form
another larval instar
When there is a
lower concentration
of juvenile hormone,
the ecdysoneinduced
molt
produces a pupa.
When ecdysone acts
in the absence of
juvenile hormone,
the imaginal discs
differentiate, and
the molt gives rise
to the adult
Insect juvenile hormones are critical developmental hormones that
have direct effects on both larval development and adult reproductive
competence. Most insect orders appear to synthesize a single JH
homolog, methyl (2E,6E)-10,11-epoxy-3,7,11-trimethyl-2,6dodecadienoate
(JH III) but Lepidoptera and at least some Diptera
synthesize additional homologs.
Although not well documented, regulation of JH production is
complicated and involves endogenous neural, neuroendocrine signals
and, in some cases, male produced exogenous regulators transferred
to the female during mating. Among virgin females, JH is required for
vitellogenesis and, thus, females do not become reproductively
competent until JH production is stimulated. To date, only two
neuropeptides that regulate JH biosynthesis in adult Lepidoptera have
been identified. These were identified e.g. from the tobacco
hornworm moth (Manduca sexta) and are:
allatotropinallatotropin
(Gly-Phe-Lys-Asn-Val-Glu-Met-Met-Thr-Ala-Arg-Gly-Phe-NH2)
allatostatinallatostatin
(pGlu-Val-Arg-Phe-Arg-Gln-Cys-Tyr-Phe-Asn-Pro-Ile-Ser-Cys-Phe-
COOH).
HormonHormonáálnlníí regulace syntregulace syntéézy feromony:zy feromony:
Coleopteran pheromone
biosynthetic pathways as
exemplified for Ips spp.
[e.g. Ips pini (Say)] and
acyclic monoterpenoid
(ipsdienol) pheromone
biosynthesis. The classical
mevalonate-based
isoprenoid pathway is
regulated by juvenile
hormone III (JH III)
at enzymatically catalyzed
steps prior to mevalonate.
Feeding on host Pinus spp.
phloem induces synthesis
of JH III by the corpora
allata.
VyskytujVyskytujíí se u hmyzu pohlavnse u hmyzu pohlavníí hormony?hormony?
EndokrinnEndokrinníí disrupce u bezobratlých:disrupce u bezobratlých:
The issue of endocrine disruption (ED) in invertebrates has
generated remarkably little interest in the past compared to
research with aquatic vertebrates in this area. However, with more
than 95% of all known species in the animal kingdom, invertebrates
constitute a very important part of the global biodiversity with key
species for the structure and function of aquatic and terrestrial
ecosystems. Despite the fact that ED in invertebrates has been
investigated on a smaller scale than in vertebrates, invertebrates
provide some of the best documented examples for deleterious
effects in wildlife populations following an exposure to endocrineactive
substances. The principal susceptibility of invertebrates to
endocrine-active compounds is demonstrated with the case studies
of tributyltin effects in mollusks and of insect growth regulators,
the latter as purposely synthesized endocrine disrupters.
The first adverse effects of TBT on mollusks were observed in
Crassostrea gigas at the Bay of Arcachon, one of the centers of
oyster aquaculture in Europe with ball-shaped shell deformations
in adults, and a dramatically decline of annual spatfall. These
effects led to a break-down of local oyster production in the bay
with marked economic consequences. Laboratory and field
analyses revealed that TBT from antifouling paints was the
causative agent with trace concentrations as low as 10 to 20 ng
TBT/L in ambient water being already effective. Another TBT
effect in molluskswas initially described in a number of regions
worldwide in the early 1970s without identifying the organotin
compound as the causative agent at that time: A virilization of
female prosobranchs, which has been termed as imposex. Imposex
is characterized by the formation of a penis and/or vas deferens
on females of gonochoristic prosobranch species and is induced at
lower concentrations than all other described TBT effects.
Furthermore, it is a specific response of organotin compounds
under field conditions. Today, imposex is known to occur in more
than 150 prosobranch species.
Imposex:Imposex:
Imposex:Imposex:
Tributyltin (TBT)
The periwinkle Littorina littorea develops a closely related
virilization phenomenon as a response to TBT exposure, termed as
intersex. Intersex females are either characterized by male
features on female pallial organs, specifically by an inhibition of the
ontogenetic closure of the pallial oviduct or female sex organs are
supplanted by the corresponding male formations particularly by a
prostate gland. Comparably to imposex, the intersex response is a
gradual transformation of the female pallial tract, which can be
described by an evolutive scheme with four stages.
Intersex development causes restrictions of the reproductive
capability of females. In stage 1, a loss of sperm during copulation is
possible and consequently the reproductive success is reduced.
Females in stages 2-4 are definitively sterile because the capsular
material is spilled into the mantle cavity (stage 2) or the glands
responsible for the formation of egg capsules are missing (stages 3
and 4). Due to female sterility, periwinkle populations can be in
decline but are not likely to become extinct because of the
planktonic veliger larvae produced by the species, as long as aqueous
TBT levels are not beyond mortality threshold concentrations for
the larvae (Matthiessen et al. 1995).
One of the most important lessons to be learnedOne of the most important lessons to be learned
from the "TBT story" and its effects in mollusksfrom the "TBT story" and its effects in mollusks
is that EDCs may impact different levels ofis that EDCs may impact different levels of
biological integrations from molecules tobiological integrations from molecules to
communities affecting also the survival ofcommunities affecting also the survival of
populations in the field. Furthermore, the casepopulations in the field. Furthermore, the case
history of TBT provides evidence forhistory of TBT provides evidence for
vertebratevertebrate--type steroids playing an importanttype steroids playing an important
functional role in a number of invertebratefunctional role in a number of invertebrate
groups, including prosobranchs.groups, including prosobranchs.
PolycyklickPolycyklickéé aromatickaromatickéé uhlovoduhlovodííky mohouky mohou
aktivovat EcR:aktivovat EcR:
PolycyklickPolycyklickéé aromatickaromatickéé uhlovoduhlovodííky mohouky mohou
aktivovat EcR:aktivovat EcR:
PolycyklickPolycyklickéé aromatickaromatickéé uhlovoduhlovodííky mohouky mohou
aktivovat EcR:aktivovat EcR:
Bisphenol ABisphenol A