S U S H M A B O P P A N A

B S C I 2 9 6 , S P R I N G 2 0 1 4

J U L I A N H I L L Y E R

Functional and Structural Characterization of Antennal APOs

in Anopheles gambiae

Mosquito Antennae

The antennae’s Johnston’s organs influence courtship and mating.

The reception of odorant and thermal cues by antennal receptor neurons controls oviposition and host-seeking behaviors.

Source: Zwiebel Lab

Flagellum

Pedicel

Scape

Antennal Accessory Pulsatile Organs (APOs)

AIMS

AIM 1: Functionally characterize the antennal APOs

AIM 2: Understand hemolymph flow within the antennal space

AIM 3: Substantiate previous studies on antennal APO structure

Location of the antennal APOs

Posterior but close to the base of the antenna (the scape)

Symmetrically located on each side of the mosquito’s dorsal midline

Found within the inside edge of the mosquito’s compound eyes

Methods: Contraction Rate Experiments

• Mosquitoes were cold anesthetized and injected with 1μm green fluorescent beads.

• Legs and wings were cut off, and mosquitoes were positioned dorsal side up.

• A 1 minute video of the APOs was acquired followed by 1 min. video of the heart.

Antennal APO Contraction Rates

The right APO contracts an average of 0.978 ± 0.23 Hz.

The left APO has an average rate of 0.983 ± 0.237 Hz.

The left and right APO contract in sync with each other.

APO Contractions Over 15 Seconds

Antennal APO vs. Heart Rates

The heart contracts, on average, at a rate of 1.78 Hz or 106.7 times a minute.

The antennal APOs contract 45% slower than the heart (paired t-test, p<0.0001).

Methods: Particle Tracking Experiments

Mosquitoes were cold anesthetized and injected with 0.5 um fluorescent beads.

Mosquitoes were restrained and positioned in the same manner as mentioned before.

Particle tracking software was used to track 6 particles/antenna: 3 up and 3 down.

Hemolymph Flow within Antennae

Particles travel up the antenna at an average velocity of 197 µm/sec and down at 51 µm/sec.

Maximum acceleration of particles going up the antenna is significantly greater when compared to downward traveling particles (t-test, p<0.0001).

Methods: Staining and Histology

Mosquito heads were fixed, dehydrated, and infiltrated by JB-4 plus.

These heads were then transferred into molding trays and allowed to polymerize

Sectioning was carried out on a JB4 Sorvall Porter-Blum microtome, and sections were loaded, 10 at a time, onto slides.

The slides were stained with Azure II Blue.

Antennal Vessel Structural Data

Sections showed clear evidence of an antennal vessel.

Antennal vessel model:

Narrow vessel allows hemolymph to be pumped up the antenna

Emptied out into the wider antennal hemocoel at the distal end of the appendage

Hemolymph returns to the head of the mosquito in a less linear manner and more slowly.

Antennal APO Structural Data

Ampulla (Am) = sac-like chamber

where hemolymph accumulates

Z-body (ZB) = dense cluster of cells attached to the contractile muscle

Passive Systole Action

Methods: FlyNap

FlyNap is a common anesthetic mixture consisting mainly of triethylamine.

Has a cardioacceleratory effect on mosquito heart rates

Method similar to contraction rate experiments using cold anesthesia.

VS.

Cold anesthesia FlyNap anesthesia

Cold vs. FlyNap Anesthesia

APO and heart contraction rates for cold anesthetized mosquitoes were significantly lower than for FlyNap anesthetized mosquitoes (t-test, p<0.0001).

Conclusions

Antennal APOs contract synchronously at an average of 0.98 Hz.

The antennal APOs contract 45% slower than the heart.

Hemolymph travels through the antenna via an antennal vessel.

The mosquito antennal APO structure was found to be in line with previous data.

FlyNap has an acceleratory effect on both APOs and the heart.

Acknowledgement: Dr. Julián Hillyer

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