Central gustatory lesions and learned taste aversions: unconditioned stimuli

Abstract

The efficacy of two different unconditioned stimuli (US) in producing conditioned taste aversion (CTA) was tested in rats after bilateral ibotenic acid (IBO) lesions of the gustatory nucleus of thalamus (TTAx) and the medial and lateral parabrachial nuclei (mPBNx, lPBNx). An initial study determined an equivalent dose for the two USs, LiCl and cyclophosphamide (CY), using non-lesioned rats. Subsequently, using a separate set of lesioned animals and their sham controls (SHAM), injections of CY were paired 3 times with one of two taste stimuli (CSs), 0.1 M NaCl for half the rats in each group, 0.2 M sucrose for the other half. After these conditioning trials, the CS was presented twice more without the US, first in a 1-bottle test, then in a 2-bottle choice with water. The acquisition and test trials had 2 intervening water-only days to assure complete rehydration. Two weeks later, the same rats were tested again for acquisition of a CTA using LiCl as the US and the opposite CS as that used during the CY trials. The SHAM and TTAx groups learned to avoid consuming the taste associated with either CY or LiCl treatment. The two PBNx groups failed to learn an aversion regardless of the US.

Fig. 1

Comparison of a conditioned taste aversion (CTA) in rats induced by the unconditioned stimuli (US) cyclophosphamide (CY) and lithium chloride (LiCl). Morning 15-min fluid intakes (mean±SEM) for the last 3 days of water baseline (Pre-ACQ) and the 14-day experimental period (Days 1 to 14). The solutions were deionized, distilled water (dH₂O) and 0.03 M Polycose, the conditioned stimulus (CS). Illness was induced by intraperitoneal (ip) injections of 0.15 M LiCl at the dose of 1.33 ml/100 g (85 mg/kg, Panel A), or 25-mg/ml CYat doses of 5, 15, and 45 mg/kg (Panels B, C, and D, respectively). The CS was followed by US injections on 3 acquisition trials (Days 1, 4, and 7). Day 10 was a one-bottle test (1-B) with the CS but no US; Day 13, a two-bottle test (2-B) with the CS, dH₂O, and no US. In each panel, open squares (□) equal intake of 1-B dH₂O during 15 min AM access; half-closed square (◨) equals intake of 2-B dH₂O. Closed-triangles (▴) equal intake of 1-B CS Polycose trials and test; semi-closed triangle (◭) equals intake of the CS in a 2-B test. (***, p<0.001).

Fig. 2

Three day extinction cycles of the conditioned taste aversions (CTA) induced by cyclophosphamide (CY) and lithium chloride (LiCl). Morning 15-min fluid intake (mean±SEM) of the 0.03 M Polycose conditioned stimulus (CS), and deionized, distilled water (dH₂O). These 3-day cycles consisted of 2 days exposure to the CS alone (1-B), then a one-day 2-bottle (2-B) choice between dH₂O and the CS. Cycles began the day after the first 2-B preference test. Closed (▲) and half-closed (◭) triangles represent the CS intake in 1-B and 2-B tests, respectively; semi-closed squares (◨) denote the intake of dH₂O in the 2-B tests (**, first significant difference in 2-B tests, p<0.01; ns, first no significant difference in 2-B tests).

Fig. 3

Photomicrographs of bilateral coronal sections from the brains of rats stained for neurons with NeuN immunohistochemistry. Panels A and B show the bilateral electrophysiologically guided ibotenic acid (IBO) lesions made at the thalamic taste area (TTAx). Panels C and D, a similar level through the TTA in a sham control rat (SHAM). Panels E and F, IBO lesions aimed at the medial parabrachial nuclei (mPBNx). Panels G and H, IBO lesions aimed at the lateral parabrachial nuclei (lPBNx). Panels I and J, the PBN in a SHAM rat. Arrows point to the target area of the lesion, black dots outline the extent of complete neuronal degeneration. Scale bar=0.5 mm. (BC, brachium conjunctivum; CP, cerebral peduncle; fr, fasiculus retroflexus; LC, locus coeruleus; lPBN, lateral parabrachial nuclei; ml, medial lemniscus; MoV, Motor neuron of trigeminal nerve; mPBN, medial parabrachial nuclei; pf, parafascicular nucleus; STA, supratrigeminal area; VPM, ventroposteromedial thalamic nuclei; 3V, third ventricle).

Fig. 4

Comparison of conditioned taste aversion (CTA) induced by the unconditioned stimuli (US) cyclophosphamide (CY) and lithium chloride (LiCl) paired with conditioned stimuli (CSs) in rats with central gustatory lesions and their sham controls. Group means (±SEM) of 15-min AM fluid intake during the last 3 days of water baseline (Pre-ACQ) and the 14-day experimental period (Days 1 to 14). On the three acquisition trials (Days 1, 4, and 7), the CS was followed by an intraperitoneal (ip) injection of the US. Day 10 was a 1-bottle (1-B) test with the CS but no US. Day 13 was a two-bottle (2-B) test with the CS, dH₂O, and no US. In the first round the US was 15-mg/kg CY (bottom panels); in the second, 85-mg/kg LiCl (top panels). The two CSs were 0.1 M NaCl and 0.2 M sucrose. They were counterbalanced across the groups and USs, but the results are collapsed here. Top panels (A–D); LiCl US. Bottom panels (E–H); cyclophosphamide US. Open squares (□) equal 1-B dH₂O intake during the 15-min AM access periods. Closed triangles (▴) equal 1-B CS intake in 1-B tests. Half-closed squares (◨) equal dH₂O intake in the 2-B test; semi-closed triangles (◭), equal CS intake in the 2-B test. (**, p<0.01; and ***, p<0.001).

Fig. 5

Comparison of afternoon water intake of rats with central gustatory lesions and their sham controls. Group means (±SEM) represent 1-h PM deionized, distilled water (dH₂O) intake during the last 3 days of water baseline (Pre-ACQ) and the 14-day experimental period (Days 1 to 14). The three acquisition trials were performed in the morning on Days 1, 4, and 7 (arrows). Day 10 was a 1-bottle (1-B) test with the CS but no US. Day 13 was a two-bottle (2-B) test with the CS, dH₂O, and no US. Closed squares (▪), LiCl; open squares (□), cyclophosphamide. (**, p<0.01; and ***, p<0.001).