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Responses of protocerebral neurons in Manduca sexta to sex-pheromone mixtures

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Abstract

Male Manduca sexta moths are attracted to a mixture of two components of the female’s sex pheromone at the natural concentration ratio. Deviation from this ratio results in reduced attraction. Projection neurons innervating prominent male-specific glomeruli in the male’s antennal lobe produce maximal synchronized spiking activity in response to synthetic mixtures of the two components centering around the natural ratio, suggesting that behaviorally effective mixture ratios are encoded by synchronous neuronal activity. We investigated the physiological activity and morphology of downstream protocerebral neurons that responded to antennal stimulation with single pheromone components and their mixtures at various concentration ratios. Among the tested neurons, only a few gave stronger responses to the mixture at the natural ratio whereas most did not distinguish among the mixtures that were tested. We also found that the population response distinguished among the two pheromone components and their mixtures, prior to the peak population response. This observation is consistent with our previous finding that synchronous firing of antennal-lobe projection neurons reaches its maximum before the firing rate reaches its peak. Moreover, the response patterns of protocerebral neurons are diverse, suggesting that the representation of olfactory stimuli at the level of protocerebrum is complex.

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Acknowledgments

The authors thank Teresa Gregory for technical assistance and Drs. Robert Mitchell, Jeffrey Riffell and Joshua Martin for valuable comments. This work was supported by NSF grant DMS-1200004 to HL and NIH grant R01-DC02751 to JGH.

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  1. Department of Neuroscience, University of Arizona, Tucson, AZ, 85721, USA

    Hong Lei, Hong-Yan Chiu & John G. Hildebrand

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  1. Hong Lei
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Correspondence to Hong Lei.

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359_2013_844_MOESM1_ESM.eps

Supplemental Figure 1. Examination of response consistency, measured by coefficient of variation (CV). The responses are quantified using Mean instantaneous firing rate within response window (a), Response delay (b) and Number of spikes within response window (c). Each matrix is consisted of the CV values (expressed as a color scale) from 34 neurons that are arranged symmetrically. The primary diagonal represents the CV values based on the multiple trials within each neuron and other parallel diagonals are based on pairwise combinations of all neurons. The mean of each diagonal is assigned to the bar graph that is placed at the top right corner. The middle section of the bar graph is indicated by double arrows and the mean ± 2 standard deviation of the bar values is indicated red and black dashed lines. (EPS 9757 kb)

359_2013_844_MOESM2_ESM.eps

Supplemental Figure 2. Dye fill reveals a partially stained protocerebral neuron (labeled as VC110804) with arborization in lateral protocerebrum (a). This neuron showed nearly identical responses to EEZ, BAL and the Mixture, as demonstrated by the peristimulus time histograms (b). The black bar below each histogram indicates olfactory stimulation. D: dorsal; V: ventral; L: lateral; M: medial; O: optic lobe; SOG: suboesophageal ganglion; EEZ: E10-E12-Z14-hexadecatrienal; BAL: bombykal, i.e. E10-Z12-hexadecadienal. (EPS 7579 kb)

359_2013_844_MOESM3_ESM.eps

Supplemental Figure 3. Two neurons were partially stained with their somata seen lateral to the ipsilateral calyx and their main neurites crossing the calyx anteriorly (arrows) toward contralateral hemisphere (a). These two neurons had sparse arborization in the ipsilateral ventral protocerebrum. The recorded neuron showed nearly identical responses to single pheromone components and their blend of different ratios (b). When the quantity of both components was raised 100 times, the response appeared to be weaker. D: dorsal; V: ventral; L: lateral; M: medial; SOG: suboesophageal ganglion; EEZ: E10-E12-Z14-hexadecatrienal; BAL: bombykal, i.e. E10-Z12-hexadecadienal. (EPS 8105 kb)

359_2013_844_MOESM4_ESM.eps

Supplemental Figure 4. A protocerebral neuron, labeled as VC100714, bilaterally connects lateral protocerebrum of both hemispheres and send information to the mushroom calyx that is ispilateral to its soma (a). The branches in the calyx display blebby processes, typical of neuronal terminals. This neuron exhibited triphasic response pattern to single pheromone components and their blend of different ratios (b). The pattern can be characterized as a short burst of spikes followed by a period of spike suppression than followed by a 2nd burst of spikes. Hisbiscus oil, containing many plant derived volatiles, evoked strong excitatory response on this neuron. D: dorsal; V: ventral; L: lateral; M: medial; SOG: suboesophageal ganglion; EEZ: E10-E12-Z14-hexadecatrienal; BAL: bombykal, i.e. E10-Z12-hexadecadienal. (EPS 9002 kb)

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Lei, H., Chiu, HY. & Hildebrand, J.G. Responses of protocerebral neurons in Manduca sexta to sex-pheromone mixtures. J Comp Physiol A 199, 997–1014 (2013). https://doi.org/10.1007/s00359-013-0844-4

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