A key goal in behavioral neurobiology is to define the cellular and molecular determinants that enable animals to decide between two or more alternative actions based on environmental cues. Thermotactic discrimination in Drosophila larvae represents a simple model to address the interplay between sensory input and choice selection. In the current study, we defined the cellular and molecular requirements for selection of slightly cool over the optimal temperature, and demonstrated how the cells and TRP channels contribute to thermotactic behavior. Specifically, we found that the chordotonal neurons functioned in the discrimination of 17.5°C and slightly cooler temperatures, and this required a TRPV channel, Iav. In support of a requirement for the chordotonal neurons, the effect due to loss of iav was reversed by introduction of the wild-type gene in chordotonal neurons. Moreover, expression of tetanus toxin in chordotonal neurons suppressed the ability to select 17.5 over 14°C, consistent with a requirement for synaptic transmission from the chordotonal neurons for cool temperature discrimination.
It has been reported previously that the terminal organs are involved in selecting 18°C over 11°C (Liu et al., 2003
). We found that the chordotonal neurons and iav
were not required for selecting the optimal temperature over temperatures cooler than 14°C. We propose that the chordotonal organ functions in the discrimination of 17.5°C versus slightly cooler temperatures (14 – 16°C), while the terminal organ functions in the selection of 17.5°C over 12°C and cooler temperatures.
The finding that Iav was required for choosing the optimal temperatures over slightly cool temperatures underscores the broad evolutionary role for TRPV channels in temperature selection. However, the requirement for Iav is distinct from mammalian TRPVs, several of which function in the responses to warm or hot temperatures (Bandell et al., 2007
; Venkatachalam and Montell, 2007
). Whether the other Drosophila
TRPV, Nan, also contributes to temperature selection is unclear, since the thermotaxis assay was complicated by the greater sedentary behavior than iav
, and the abnormally high turning even at 17.5°C. As a result, the nan
larvae traveled small distances and remained near the initial site in which they were placed. Nevertheless, as is the case for Iav, the Nan channel is also expressed in chordotonal neurons, and therefore could potentially function in cool sensation (Gong et al., 2004
). However, at least one other TRP channel, Pain, is expressed in chordotonal neurons (Tracey et al., 2003
), but is not required for choosing 17.5°C over mildly cool temperatures.
In addition to Iav, another TRP channel, TRPL, functions in cool sensation (Rosenzweig et al., 2008
). As was previously reported for TRPL (Rosenzweig et al., 2008
), expression of Iav in oocytes did not elicit a cool-activated current (data not shown). Thus, it is not clear whether activation of either of these channels by thermal cool is direct or indirect. In contrast to iav
, we did not detect a requirement for trp
in temperature discrimination in the cool range, which differs from a previous report (Rosenzweig et al., 2008
). However, a role for trp
was less clear, given the lack of phenotype when the trp
mutation was placed in trans
with a deficiency, which uncovered the gene (Rosenzweig et al., 2008
). Nevertheless, given that the previous and current studies focused on early and late stage larvae respectively, we cannot exclude that there are developmental differences in requirements for the TRP channel for sensing mildly cool temperatures.
Even though wild-type third instar larvae prefer 17.5°C over any other temperature, we found that the P.I. increased significantly in proportion to the alternative temperature (~0.2, 12°C; ~0.35, 14°C; ~0.7, 24°C). We suggest that these differences are only partially due to the slower movements at the lower temperatures. The variations in the average P.I. values may reflect differences in the molecules, mechanisms and cell types involved in discriminating the optimal temperature (17.5°C) from mildly cool (14–16°C), cool (12°C) and comfortably warm (22–24°C) temperatures. In support of this proposal, inhibition of neurotransmission in chordotonal neurons specifically impaired the ability to discriminate between the preferred temperature over 14°C (), and did not reduce selection of 17.5° over either 12°C or 24°C. Furthermore, a Gq/phospholipase C/TRPA1 thermosensory signaling cascade participates in choosing the optimal temperature over other temperatures in the comfortable range (20–24°C) (Kwon et al., 2008
), while Iav and TRPL are required for opting for 17.5°C over mildly cool temperatures.
A critical question is how the thermotactic selection of the optimal over mildly cooler temperatures is accomplished via the chordotonal neurons and iav. The following three observations lead us to propose a model. First, iav mutant animals retain the ability to select the optimal temperature over very cool and comfortably warm temperatures. These results argue against a requirement for iav for positive selection of 17.5°C only, since this optimal temperature is still selected versus some temperatures. Second, stimulation of iav-expressing neurons with channelrhodopsin induces an avoidance response to light. Third, the modestly cooler temperature increased the average turning angles and the total number of turns (θ ≥ 45°) relative to wild type. Thus, we conclude that iav-dependent decision to choose the optimal over slightly cooler temperatures is mediated by increased activity of chordotonal neurons, which stimulates an avoidance response by increasing the number and magnitude of turning angles.