Food enhances taste responses in ASH neurons. (A) Avoidance of soluble repellents is increased by food in wild-type animals. Fraction of animals on- and off-food reversing after delivery of a drop with repellent. Avoidance to 2 mM CuCl₂, 10 mM primaquine and 0.5 M glycerol is increased on-food. (^***P⩽0.0001, t-test), N⩾68. (B, C) Average neuronal responses in ASH. Grey shading indicates the duration of the stimulus. (B) Neuronal responses to 2 mM CuCl₂ are increased by food. (C) Neuronal responses to 0.5 M glycerol are increased by food. (D) Quantification of ASH responses to 2 mM CuCl₂ and 0.5 M glycerol on- and off-food. Blue circles represent individual animals assayed. Black diamonds indicate average value. Food increases the magnitude of ratio changes in wild-type animals. Statistical significance according to the Mann-Whitney test is indicated (^**P<0.01), N⩾15.

Food modulation of ASH involves dopamine. (A) Avoidance is increased by exogenous dopamine (DA). Avoidance is not increased by exogenous serotonin. Fraction of wild-type animals off-food, on-food, on 10 mM DA and on 10 mM serotonin, reversing after delivery of a drop with CuCl₂. (^***P<0.0001 ratio t-test), N⩾170. (B) Avoidance is not increased by food but by exogenous DA in cat-2(e1112). Fraction of wild-type and cat-2(e1112) animals, on- and off–food, and on and off 10 mM DA, reversing after delivery of a drop with 2 mM CuCl₂. N⩾213. (C–F) Average neuronal responses in ASH. Grey shading indicates the duration of the stimulus. (C) Neuronal responses to 2 mM CuCl₂ are increased by food in wild-type animals. (D) Responses to 2 mM CuCl₂ in wild-type animals are increased by 10 mM exogenous DA. (E) Neuronal responses to 2 mM CuCl₂ are not affected in cat-2(e1112). (F) Neuronal responses to 0.5 M glycerol are not affected in cat-2(e1112). (G) Responses to nose touch are not affected by 10 mM serotonin and 10 mM DA in wild-type animals. (H) Quantification of ASH responses. Blue circles represent individual animals assayed. Black diamonds indicate average value. Exogenous DA increases the magnitude of the ratio change in wild-type animals. In cat-2(e1112), ASH responses to 2 mM CuCl₂ and 0.5 M glycerol are not increased by food. Exogenous DA or serotonin does not affect the ratio change in wild-type animals in response to nose touch. (^***P<0.001, ^**P<0.01, Mann–Whitney rank sum test), N⩾12.

Food and dopamine do not affect reversals caused by optogenetic activation of the ASH neurons. (A–D) Reversals following optogenetic activation of the ASH neurons. (A) Reversal probability is not increased by food. (B) Reversal duration is not increased by food. (C) Reversal probability is not increased by DA. (D) Reversal duration is not increased by DA.

Food and dopamine inhibit adaptation to repellents. (A, B) Fraction of animals responding to repeated drops of 10 mM CuCl₂. (A) Wild-type animals adapt slower on-food and 10 mM DA (P⩽0.036, t-test), N⩾30. (B) cat-2(e1112) animals do not adapt slower on-food. cat-2(e1112) adapts faster than wild type on-food, dop-4(ok1321) is not significantly different from wild-type animals. (^***P<0.001, ^**P<0.01, ^*P<0.05, t-test), N⩾37. (C,D) Fraction of animals responding before and after a chronic stimulation. (C) cat-2(e1112) and dop-4(ok1321) adapt like wild type to chronic stimulation off-food. (D) cat-2(e1112) adapts faster than wild type on-food. dop-4(ok1321) is not significantly different from wild type. (E–G) Average neuronal responses in ASH before and after chronic treatment with 10 mM CuCl₂. Grey shading indicates the duration of the stimulus. (E) Wild-type and cat-2(e1112) animals adapt completely off-food, and show no post-chronic responses. (F) On-food, wild type does not fully adapt, and still displays post-chronic responses. cat-2(e1112) completely adapts on-food. (G) Wild type does not fully adapt and displays post-chronic responses in exogenous DA. (P<0.001, t-test), N⩾24. (H) Quantification of ASH responses before and after chronic treatment. Blue circles represent individual animals assayed. Black diamonds indicate average value. Wild type adapts completely to a chronic stimulation with 10 mM CuCl2 off-food, and adapts less on-food or 10 mM DA. (P<0.01, Mann–Whitney rank sum test), N⩾17.

Dopaminergic neurons with intact cilia are required for increased avoidance on food. Fraction of worms reversing on- and off-food after a drop of 0.5 M glycerol. daf-12(sa204) animals show increased responses on food, whereas cilia-defective daf-19(m86);daf-12(sa204) animals exhibit defective avoidance on- and off-food. Rescuing DAF-19c under the gpa-11 (ASH, ADL) or gpa-13 (ASH, ADF, AWC, PHA and PHB) promoters results in normal responses off-food, but does not increase avoidance on food. Rescuing DAF-19c using both the gpa-13 and dat-1 (dopaminergic neurons) promoters leads to increased avoidance on food. (^***P<0.001, t-test), N⩾103.

Optogenetic activation of the dopaminergic neurons induces slowing and increases avoidance. (A) Normalized speed in animals expressing ChR2 under the dat-1 promoter (dopaminergic neurons). Shown is the normalized speed measured every 10 s. Animals not grown on retinal are not affected by blue-light stimulation. In animals grown on retinal, a 10-s-long blue-light stimulation induces slowing, which lasts for 60 s. (P⩽0.0006, t-test), N⩾50. Blue-light stimulation does not induce slowing in cat-2(e1112). dop-4(ok1321) slows like wild type. (B) Fraction of animals reversing after blue-light stimulation, followed by a drop of 2 mM CuCl₂. Time between light activation and drop delivery is indicated. Animals not grown on retinal are not affected by the blue-light stimulation. Animals expressing ChR2 under the tph-1 promoter (serotonergic neurons) and grown on retinal do not have increased avoidance after the light flash. Animals expressing ChR2 under the dat-1 promoter and grown on retinal show increased avoidance after blue-light activation, and this effect lasts 1–2 min after the light flash. (P⩽0.0002, t-test), N⩾137. Avoidance is not affected in dat-1∷ChR2 transgenes in cat-2(e1112) and dop-4(ok1321) backgrounds. (C) Food does not further enhance avoidance in animals expressing ChR2 under the dat-1 promoter. Fraction of animals reversing after blue-light stimulation, followed by a drop of 2 mM CuCl₂. dat-1∷ChR2; lite-1(ce314) animals not grown on retinal have enhanced responses on food. dat-1∷ChR2; lite-1(ce314) animals grown on retinal have enhanced responses both off- and on-food, and the avoidance is not further increased in the presence of food. (^***P<0.001, t-test), N⩾150.

The dopamine receptor DOP-4 mediates the effect of food on avoidance. (A) Avoidance is not increased by food in dop-4(ok1321). Fraction of animals responding to 2 mM CuCl₂. (^***P<0.001, t-test), N⩾150. (B) Avoidance in dop-4(ok1321) is not increased by exogenous DA. 10 mM exogenous DA increases the avoidance to 2 mM CuCl₂ in wild type and cat-2(e1112), but not in dop-4(ok1321). (^***P<0.001, t-test), N⩾113. (C,D) Average neuronal responses in ASH. Grey shading indicates the duration of the stimulus. (C) Magnitude of ASH responses to 2 mM CuCl₂ is not increased by food or exogenous DA in dop-4(ok1321). In wild-type animals, the magnitude of responses to 2 mM CuCl₂ is increased by food and 10 mM exogenous DA, but not in dop-4(ok1321). Expression of genomic dop-4 DNA under the sra-6 promoter (ASH, ASI, PVQ) rescues ASH responses in dop-4(ok1321). Shown are the average traces. (D) Magnitude of ASH responses to 0.5 M glycerol is not increased by food in dop-4(ok1321). In wild-type animals, the magnitude of responses is increased by food, but not in dop-4(ok1321). (E, F) Quantification of ASH responses. Blue circles represent individual animals assayed. Black diamonds indicate average value. (^***P<0.001, ^**P<0.01, Mann-Whitney rank sum test) (E) Wild-type animals have increased responses to 2 mM CuCl₂ and 0.5 M glycerol on exogenous DA. In dop-4(ok1321), the responses are not increased. (F) Rescue of dop-4(ok1321) under the sra-6 promoter restores the magnitude of the response to 10 mM DA. (^***P<0.001, ^**P<0.01, Mann–Whitney rank sum test), N⩾9.

The dopamine receptor DOP-4 functions in ASH to modulate avoidance behaviours. (A) Transgenic rescue of dop-4(ok1321). Fraction of animals responding to 2 mM CuCl₂. Expressing genomic dop-4 DNA under its own promoter (ASG, AVL, CAN and PQR), or the sra-6 (ASH, ASI and PVQ) or gpa-13 (ASH, ADL, AWC, PHA and PHB) promoters restores the avoidance on food. Expression under the gcy-5 promoter (ASER) does not rescue. (^***P<0.001, t-test), N⩾126. (B) RNAi knockdown of DOP-4 in ASH results in defect avoidance on food. Fraction of animals responding to 2 mM CuCl₂. Animals expressing dop-4 RNAi under its own promoter, or under the sra-6 and gpa-13 promoters, both of which express in ASH, do not have increased avoidance on food. Animals expressing dop-4 RNAi under the gcy-5 and flp-17 promoters, which do not express in ASH, display wild-type behaviour. (^***P<0.001, t-test), N⩾94.