Figure 6.

Third-order neurons extract distinct features of a male from cVA

(A) Circuit diagram of the first three layers of cVA processing based on hemibrain. Number of synaptic connections: ORN-lvPN: 286; lvPN-pC1: 45; ORN-lPN: 9,187; lPN-AV2a2: 115; lPN-aSP-g: 37. See also Tables S3 and S4.

(B) Top: pC1 axons respond to lvPN optogenetic activation. Red area: optogenetic stimulation; average response from 6 flies, 6 stimulations, and shaded area is SEM. Bottom: EM reconstruction of lvPNs (red) and the five pC1 cells (a–e) (yellow, from Wang et al.³⁴) neurons in FAFB, top view. Red circles: location of lvPN to pC1 synapses. Gray square: imaging ROI for pC1 recordings. Scale bars, 20 μm.

(C and D) Left: EM reconstructions in FAFB of pC1 (C, from Wang et al.³⁴) and AV2a2 (D) neurons. Scale bars, 20 μm. Right: GCaMP6f responses in pC1 (C) and AV2a2 axons (D) to male presentation, same stimulus as Figure 2G. Average responses from 6 flies, 6 trials, and shaded area is SEM of biological replicates.

(E) GCaMP6f responses in ORN axons to male presentation (0.75 mm) at different speeds (shown on the left). Average responses from 11 flies, 6 trials, and shaded area is SEM of biological replicates. Black trace: male position, dashed line: end of the approach.

(F) GCaMP6f responses in lPN axons, stimulus as in (D), n = 7, 6 trials. Data with the highest speed was included in Figure 2H.

(G) GCaMP6f responses in AV2a2 axons, stimulus as in (D), n = 6, 6 trials. Data with the highest speed also shown in (C).

(H) The differential of the lPN response trace to presenting a male at different speeds, based on data in (E).

(I) Peak responses at different male speeds on ORNs, lPNs, and AV2a2, based on data in (D)–(F). Data points from individual flies connected with gray lines.

(J) lPN response differential peaks correlate with AV2a2 peak responses. Error bars are standard deviation. Dashed line shows the linear fit, R² = 0.71, p = 0.36.

(K) Circuit diagram of AD1g1 input cell types. Number of synaptic connections: AV2a2-AD1g1: 142; LPLC2-AD1g1: 474; LC4-AD1g1: 434.

(L and M) Manipulating AV2a2-SS in females paired with wild-type males.

(L) Pulsed optogenetic activation had no effect on female receptivity.

(M) Blocking with Kir2.1 decreased female receptivity.

See also Figure S6.