Sem ttulo de diapositivo

1
Functional Asymmetry in the Olfactory System of a
Flatfish, the Senegalese Sole (Solea senegalensis)
Zélia Velez1,2,3, Peter C. Hubbard1, Kevin J.
Weham3, Eduardo N. Barata1,2, Jörg D. Hardege3
and Adelino V.M. Canário1
1. Centro de Ciências do Mar, Faro, Portugal 2.
Departamento de Biologia, Universidade de Évora,
Portugal 3. University of Hull, UK
2
The Selenagese Sole (Solea senegalensis)
High commercial value increased aquaculture
effort. Flatfish left (lower) nostril does not
migrate. Functional asymmetry in the olfactory
system? Lower epithelium prey
location/identification Upper epithelium
chemical communication
3
Methods
Electro-olfactogram (EOG) recorded from the
olfactory epithelium Olfactory potency
identification of odorants Cross-adaptation
receptor specificity Pharmacological inhibition
- transduction pathways Chromatography of
biological fluids Solid-phase extraction
crude separation of odorants HPLC fine
separation and purification of odorants LC-MS
identification of odorants
4
Methods
Food- and conspecific-related odorants
Bile and intestinal fluid Ragworm-conditioned
water
Total EOG
C-18 Sep-Pak
HPLC and EOG
Filtrate
Eluate
EOG
The ragworm Hediste diversicolor is one of the
soles main prey. They live buried in the sand or
mud (of estuaries). Bile acids are potent
odorants in fish.
5
Methods
The electro-olfactogram (EOG)
Velez et al. (2005) Physiol. Biochem. Zool. 78
756-765
EOG is a spacio-temporal summation of generator
potentials of olfactory receptor neurones.
L-cysteine is detected with equal sensitivity by
the two olfactory epithelia responses are
normalised to that of 10-3 M L-cysteine.
6
Results I
HPLC fractionation of ragworm-conditioned water
Fractions 1 and 2 contain the odorants of
interest active odorant identified using LC-MS
7
Results I
Identification of ragworm-related odorants
1-methyl-L-tryptophan
Active odorant identified as 1-methyl-L-tryptophan
. Ragworms release this amino acid, along with
L-phenylalanine and L-glutamine
8
Results I
Olfactory sensitivity
Cross-adaptation - selectivity
Lower epithelium has greater sensitivity to
1-methyl-L-tryptophan.
9
Results I
Olfactory sensitivity to conspecific odorants
Bile
Intestinal fluid
Mucus
Velez et al. (2007) Gen. Comp. Endocrinol. 153
418-425
Conspecific body fluids are consistently better
detected by the upper olfactory epithelium
10
Results I
Chromatographic separation of odorants in
intestinal fluid
Velez et al. (2009) J. Comp. Physiol. A 195
691-698
Bile acids contribute about 40 of the olfactory
potency of intestinal fluid. Bile acids in
fraction 4 were identified by LC-MS.
11
Results I
Identification of odorants in intestinal fluid
Taurocholic acid
Taurolithocholic acid
Velez et al. (2009) J. Comp. Physiol. A 195
691-698
Two major bile acids identified in the intestinal
fluid (and bile fluid). A third, minor, bile acid
was not identified.
12
Results I
Olfactory sensitivity to bile acids
Taurocholic acid
Taurolithocholic acid
Sole release taurocholic acid this is detected
better by the upper olfactory epithelium
13
Conclusions I
Prey-related odorants including
1-methyl-L-tryptophan are better detected by
the lower olfactory epithelium. Conspecific-relat
ed odorants including taurocholic acid are
better detected by the upper olfactory
epithelium. There is functional asymmetry in the
olfactory system of this flatfish. Are the
receptors and transduction pathways different?
14
Results II Cross-adaptation
1-methyl-L-tryptophan
Lower epithelium has greater specificity for the
ragworm odorant 1-methyl-L-tryptophan.
15
Results II Cross-adaptation
taurocholic acid
Upper epithelium has greater specificity for
taurocholic acid.
16
Results II Transduction
U73122 PLC inhibitor SQ-22536 AC inhibitor
L-cysteine
L-cysteine
L-cysteine is detected equally by the two
epithelia The effects of the two drugs are the
same in the two epithlia The transduction pathway
for L-cysteine is mainly via PLC
17
Results II Transduction
1-methyl-L-tryptophan
U73122 PLC SQ-22536 AC
The transduction pathway for 1-methyl-L-tryptophan
is mainly PLC mediated There are differences
between the two epithelia
18
Results II Transduction
taurocholic acid
U73122 PLC SQ-22536 AC
Both phospholipase C and adenylate cyclase are
involved in transduction of taurocholic
acid There are differences in the two epithelia
19
Conclusions II
There are specific receptors for
1-methyl-L-tryptophan in the lower
epithelium. There are specific receptors for
taurocholic acid in the upper epithelium. The
transduction pathway for 1-methyl-L-tryptophan is
mainly phospholipase C mediated. The
transduction pathway(s) for taurocholic acid
is/are mediated by both phospholipase C and
adenylate cyclase.
20
Summary
Specific receptors in the lower olfacotry
epithelium for prey-related odorants
(1-methyl-L-tryptophan) Specific receptors in
the upper olfactory epithelium for
conspecific-related odorants (taurocholic
acid) This suggests specialisation of the two
epithelia functional asymmetry Does the
processing of the olfactory information in the
olfactory bulb (and beyond) also show asymmetry?
21
The End
Thank you for your attention
Funded by FCT grants SFRH/BD/16242/2004 and
POCI/BIA-BMC/55467/2004