Sunday, June 12, 2011

A Walk Along the Paper Trail: The Carlson Lodge, Part I

One of the big open questions in olfaction research is how odorant receptors bind to and respond to different odorants.  In mammals, this question is complicated by the sheer number of olfactory receptors expressed (~400 in humans), but the issue is much simpler in Drosophila, which express only 60 olfactory receptors.  One of the leading labs studying the relationship between olfactory receptor genes, receptors, and behaviour is the Carlson lab at Yale (as a tyro to olfaction, I didn't know John Carlson's name, but damn that's a good publication record).  They recently published a paper looking at how a small set of odorants, pyrazines, are represented in the olfactory receptor neurons of drosophila larva. Since I don't know much about odorant receptors or fly olfaction, I decided to take a walk down the paper trail, and see what I could see.

Olfactory receptors, their neurons, and empty neurons

The first paper relevant to our trip is from 2003, and describes the expression of  two odorant receptors in adult flies, Or22a and Or22b.  Using in situ hybridization and a Gal4-UAS-GFP system, Dobritsa found that both of these receptors were expressed in a subset of basiconic sensilla (not that I know what a basiconic sensilla is), and eventually narrowed their expression down to a neuron labeled "ab3A."  They presented a number of odors, and confirmed that the Or22a/b expressing neurons had the same odor response profile as ab3A neurons.

While Or22a is expressed in ab3A neurons, it does not necessarily mean that ab3A's odorant response is solely determined by Or22a or Or22b.  To investigate that, they knocked out Or22a and/or b, and found that only Or22a was necessary for olfactory responses.  Then to see whether a single Or is sufficient to drive ORN responses, they expressed another olfactory receptor Or47a under the Or22a-Gal4 driver.  When they did this, they found that the ab3A neuron now had the response profile of Or47a neurons (identified as ab5B). 

Or22a neurons expressing Or47a (UAS-47a) have the same response profile as ab5B neurons, different from the ab3A neurons it typically shares a profile with. From Dobritsa et al, 2003.
The Carlson lab continued to use this expression system in the next three papers, dubbing it the "empty neuron" system. In the final section of the paper, they showed that the Or22a-driven UAS-47a neurons still mapped to the same glomerulus, DM2, as Or22a neurons.

Sitting here in 2011, it is easy to take many of the conclusions of this paper for granted, and I don't know the history of odorant receptor or drosophila olfaction research well enough to put this in historical context.  Of course ORN responses are determined by the Or they express.  There are some cute findings here.  The Or expression was sex-dependent.  And the fact that both Or22a/b are expressed in the same neuron, but only Or22a is important is curious.

The most intriguing part, to my naive view, is that last bit, where the Or22a-Gal4/UAS-Or47a neurons still map to the same place as Or22a neurons.  They mention a couple explanations: there could be other guiding odorant receptors; mammals continually regerenate ORNs, while flies don't, so the map could be established before Or22a expression; the mammalian system has more receptors, and is more complex.  This is something I should follow up...

Larva ORs differ from adult ORs

While the previous paper focused on adult flies, this next paper focused on drosophila larva.  Kreher et al performed RT-PCR on larval RNA, and identified 23 Or genes that were expressed.  Thirteen of them were the same genes as expressed in adults, while ten of them had not been detected in adult flies. Then using Or-Gal4 and UAS-GFP, they showed that each Or was expressed in one neuron.

To characterize the odorant receptive field of these newly identified Ors, they used the empty-neuron system developed by Dobritsa, and expressed the Ors in adult flies.  They tested twelve of these Ors, and found that 11 of them responded to odors (pardon the synecdoche of saying Ors respond, it's the ORNs expressing them that respond).  Some of the Ors responded to one odorant, while others responded to a variety.

Some Ors respond to multiple odorants, while others are narrowly tuned (within this set of stimuli).
From Kreher et al, 2005.
They also characterized the temporal dynamics of these responses, and showed that they can be inhibitory, delayed, transient, or sustained.  They also classified the Ors into aliphatic and aromatic groups, and claimed there was a glomerular map, but that seemed fishy to me.

Some responses are inhibitory (top left), transient (top right), or delayed (bottom right).
From Kreher et al, 2005.
The main point of this paper, identifying unique Ors expressed in larva, is fairly straightforward.  What's interesting to me, as a wannabe systems neuroscientist, are the temporal dynamics of the ORN responses.  They had previously characterized similar responses in adults, but the idea of an inhibitory response, is new to me. In the discussion, they guess that the OR is tonically moderately active, and normally is stabilized in an active state by the odor (reviewed here).  Then the inhibitory odors are able to suppress firing be turning off the active state.  Alternatively, the OR might signal to multiple G-proteins, depending on its conformation (they did not consider this possibility, I wonder why).  Since this is five years old, I bet someone has figured this out by now.

And that's where I'm going to leave it for now.  Next post I'm going to cover Keher's 2008 paper (which is pretty awesome), and the paper that made me start this journey, Montague 2011.

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