Chemicals released by male sea cucumber mediate aggregation and spawning behaviours

Abstract

The importance of chemical communication in reproduction has been demonstrated in many marine broadcast spawners. However, little is known about the use of chemical communication by echinoderms, the nature of the compounds involved and their mechanism(s) of action. Here, the hypothesis that the sea cucumber Holothuria arguinensis uses chemical communication for aggregation and spawning was tested. Water conditioned by males, but not females, attracted both males and females; gonad homogenates and coelomic fluid had no effect on attraction. Male spawning water, but not female spawning water, stimulated males and females to release their gametes; the spermatozoa alone did not induce spawning. H. arguinensis male spawning water also induced spawning in the phylogenetically related H. mammata. This indicates that males release pheromones together with their gametes that induce spawning in conspecifics and possibly sympatric species. Finally, the male pheromone seems to be a mixture with at least one labile compound (biological activity is lost after four hours at ambient temperature) possibly including phosphatidylcholines. The identification of pheromones in sea cucumbers offers a new ecological perspective and may have practical applications for their aquaculture.

Figure 1

Schematic representation of the experimental setup used in the attraction experiment. (a) Overhead view showing the entry area A, the stimulus and control sides B or C. (b) Side view showing the position of the camera IR-C and light source IR-L. SC indicates the initial positioning of the test sea cucumber.

Figure 2

Percentage of first choice (a,b) and relative time (c,d) spent in the conditioned-conspecific water (stimulus) and in the control arm (sea water). M corresponds to male receiver and F to female receiver. Error bars are one standard deviation. *p < 0.05, **p < 0.01.

Figure 3

Percentage of first choice (a,b) and relative time (c,d) spent in the coelomic fluid (stimulus) and in the control arm (sea water). M corresponds to male receiver and F to female receiver. Error bars are one standard deviation.

Figure 4

Percentage of first choice (a,b) and relative time (c,d) spent in the gonad (ovary and testis) homogenate (stimulus) and in the control arm (sea water). M corresponds to male receiver and F to female receiver. Error bars are one standard deviation.

Figure 5

(a,b) LC-MS single ion traces of methanol extracts at m/z 808.7 under positive polarity. (a) Male before (blue) and after (red) spawning; (b) female before (blue) and after (red) spawning. (c,d) Fragmentation spectra. (c) MS2(808.7); (d) MS2(790.5), the standard 1,2-stearoyl phosphatidylcholine. The inserted structures were assigned to ion with m/z 184.1. The arrows indicate the fragmented peaks.

Figure 6

(a,b) LC-MS single ion traces of methanol extracts at m/z 287 under positive polarity. (a) Male before (blue) and after (red) spawning; (b) female before (blue) and after (red) spawning. (c–e) Fragmentation spectra of m/z 287. (c) MS2(287); (d) MS3(287→245); (e) MS4(287→245→203). The arrows indicate the fragmented peaks.