Conditional disruption of synaptic transmission induces male-male courtship behavior in Drosophila

Abstract

It is reported here that male-male courtship behavior is evoked instantaneously in the fruit fly Drosophila by conditional disruption of synaptic transmission. A temperature-sensitive allele of the Drosophila dynamin gene shibire (shi(ts1)) was expressed by using the GAL4/UAS system to disrupt synaptic transmission from GAL4-positive neurons in a temperature-dependent manner. An enhancer-trap GAL4 line C309 directing shi(ts1) expression in central and peripheral neurons (C309/UAS-shi(ts1)) initiated stereotypical precopulatory behavior toward other mature males immediately after a temperature shift from the permissive to restrictive temperature. At the restrictive temperature, C309/UAS-shi(ts1) males formed "courtship chains" and exhibited abnormally high levels of head-to-head interactions. The temperature-induced male-male courtship is attributable not to an increase in sexual attractiveness but to an increase in sexual activity of C309/UAS-shi(ts1) males. Interestingly, the temperature-induced increase in sexual activity is specific toward male partners, because C309/UAS-shi(ts1) males courted receptive virgin females less vigorously and copulated less efficiently after shifted to the restrictive temperature. Among the GAL4-positive neurons in C309, conditional disruption of certain cholinergic neurons but not the mushroom body intrinsic neurons plays a critical role in the induction of male-male courtship. These neurons may be involved in inhibitory systems that normally suppress aberrant male-male courtship. The presented strategy that can induce behavioral abnormalities by disrupting synaptic transmission in an acute and noninvasive manner will allow further exploration as to how distinct neuronal groups control sexual orientation and other aspects of reproductive behavior in Drosophila.

Figure 1

GAL4 expression patterns of line C309 visualized by a GFP reporter gene. Distinct expression is observed in widespread regions of the adult brain (frontal view, A; posterior view, B) including α, β, and γ lobes of the MBs, the cell body layer of Kenyon cells (k), the pars intecerebralis (p), the AL (al), the optic lobes (ol), and the suboesophageal ganglion (sog). GFP expression is seen also in restricted sensory neurons in the labial palps (lp) (C), the tarsus (ta) (D), and the femur (fe)–trochanter (tr) junctions (E) of legs, the proximal radius of wings near the humeral cross-vein (hcv) (F), the first and third longitudinal wing veins (G and H), and the clasper (cl) of the male external genitalia (I) but not in the maxillary palp (mp) (C). [Scale bars: A, 50 μm (applied to B and C); D, 25 μm (applied to E–I).]

Figure 2

High levels of intermale interactions in populations of C309/UAS-shi^ts1 males evoked by a temperature shift from 19 to 30°C. C309/UAS-shi^ts1 males display a stereotyped sequence of actions toward other males including tapping (A, arrow), unilateral wing vibration (B, arrow), and licking and abdominal curling (C, arrow and asterisk, respectively). Note that the courted male in A is flicking both of his wings as a possible rejection response (A, asterisk). A group of C309/UAS-shi^ts1 males courting one another form a courtship chain (D) and ring (E). Arrows indicate flies exhibiting unilateral wing vibration. Shown are head-to-head interactions observed in populations of C309/UAS-shi^ts1 males, in which a male taps at another male's head (F, arrow) and extends one (G, arrow) or both (H, asterisk) of his wings.

Figure 3

Quantitative analysis of courtship chaining behavior and head-to-head interactions. Typical intermale interactions among five flies of C309/UAS-shi^ts1 at 30°C. (Upper Left) A courtship chain composed of five males. (Upper Center) Four males showing courtship behavior and a male fly flicking both wings. (Upper Right) Two pairs of males exhibiting head-to-head interactions (asterisks). The average (±SEM) of chaining index (Lower Left) and head-to-head interaction index (Lower Right), which are determined for indicated genotypes of male flies at either 30 or 19°C (see Materials and Methods).

Figure 4

Courtship index for isolated male–male (Left) or male–female (Right) pairs. The average (±SEM) of indicated number of measurements with different pairs is shown.

Figure 5

Mating success rate of single male and female pairs. The percentage of pairs that have copulated in the 1-h observation period is shown. The number of independent pairs observed for each male–female combination is indicated.

Figure 6

GAL4 expression patterns in the adult brain of different enhancer trap GAL4 lines visualized by a GFP reporter gene. Lines C747, OK107, c492b, and 17d show prominent GFP expression in the MB intrinsic neurons. The introduction of a Cha^3.3kb-GAL80 construct into the C309/UAS-GFP flies suppresses GFP expression in the most part of the brain, whereas the pars intecerebralis (p), the MB α/β lobes, and neurons localized in the lateral and medial regions of the suboesophageal ganglion (sogl and sogm, respectively) remain GFP-positive.