CA can be dissolved in HydroGel instead of water. If bottled CA water cannot easily be provided (during travel, due to cage size restrictions, or when head implants preclude the use of bottle-top cages), CA can be mixed into HydroGel cups (https://www.clearh2o.com/product/hydrogel/) as an alternative to liquid water. HydroGel was melted by placing unopened 56-g cups in a 60°C oven until the gel had liquified (3–5 h). CA powder was then mixed into the liquefied gel and stirred thoroughly, before resealing the HydroGel cups and letting them solidify at 4°C. As the flavor and perceived aversiveness of CA may differ when dissolved in water or HydroGel, we again titrated CA concentrations to achieve stable animal weights. The observation that higher concentrations of CA are required in HydroGel to achieve the same behavioral effects agreed with informal human flavor perception of both substances. A, In a first cohort of animals (cohort 5: n = 5, 15 months), this required increasing concentrations from 1% to 6% m/v. Weight (from baseline on daily measured water), as animals were given free HydroGel with different concentrations of CA. B, With a second cohort of animals (cohort 6: n = 10, 15–16 months), switching from plain HydroGel to 6% CA HydroGel resulted in weights close to the institutional minimum of 80%. At 5% CA HydroGel, all animals showed stable weights. C, In our cohort of trained animals (Fig. 2), we confirmed that a weekend regime of 2% CA could be replaced by 5% CA in HydroGel. As in Figure 2, but comparing 2% CA water with 2% CA HydroGel and 5% CA HydroGel on weekends (same animals as shown in Fig. 2; see also Extended Data Fig. 2-1). Neither weekly trial counts (t(5) = –0.932, p = 0.3941, Bf10 = 0.522) nor performance on easy trials (t(5) = –1.444, p = 0.2083, Bf10 = 0.771) were significantly different in weeks following 2% CA water versus 5% CA HydroGel. Download Figure 2-2, TIF file.

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