Molecular correlates of time-of-day specific changes in olfactory sensitivity and behavior in Anopheles gambiae

Insect physiology and behavior are regulated on a 24 hr basis by an endogenous circadian clock and by the light:dark (LD) cycle. In Anopheles gambiae, the major malaria vector, daily rhythms in flight activity, mating, sugar- and blood-feeding, and egg lying are reported. Such behaviors are further modulated by olfaction. The objective of the current study was to test for temporal changes in mosquito olfactory sensitivity and to examine the underlying molecular mechanism.

Earlier DNA microarray analysis revealed a strong rhythmicity in a large set of genes, including Odorant Binding Proteins (OBPs) at the mRNA level (Rund et al., 2011 PNAS 108: E421-E430; www.nd.edu/~bioclock). Here, we report the results from detailed electrophysiological and behavioral analyses undertaken in host-seeking female An. gambiae under LD conditions and challenged with major host derived stimuli. Mosquitoes were tested for dose-dependent responses with different compounds, and at four times of the 24 hr day, corresponding to day, dusk and night phases of the LD cycle. EAG data revealed time-of-day specific differences in olfactory sensitivity. Specifically, significant differences were observed for the major chemostimuli such as indole, nonanal and geranyl acetone that require OBPs for the sensitive detection, but not for hexanoic acid, a highly soluble stimulus that does not require OBPs. For indole, nonanal and geranyl acetone, peak sensitivity was found to occur at Zeitgeber time (ZT) 16 (night), and the lowest sensitivity at ZT8 (day) (ZT12 is time of lights off on 12:12 LD cycle).

To understand the molecular basis for these differences in olfactory sensitivity, we measured major chemosensory proteins by quantitative mass spectrometry in time-specific collections of female antennae or whole head appendages. We find that the electrophysiological responses correlate with diel changes in protein abundance of OBPs. The pre-dusk/dusk peak in OBP gene expression corresponds with peak protein concentrations, increased olfactory sensitivity and enhanced biting behavior, each occurring during the night.