COMPONENT 6 > 2008 Progress Report Summary

We have made remarkable progress during the past year, and have completed three generations of selection of both the high and low methamphetamine (MA) drinking (high=MAHDR, low=MALDR) and high and low sensitization (high=MAHSENS, low=MALSENS) lines (Specific Aim 1). In both cases, response to selection indicated the importance of genetic influences on these traits. These selections were completed in coordination with the MARC animal core, and the animal core generated additional animals for the correlated response testing described below.

Progress toward creation of short-term selected lines

For the MADR lines, mice were phenotyped for 2-bottle choice of 20 mg/L then 40 mg/L MA versus tap water, with each MA concentration offered for a 4-day period. Average mg MA/kg consumed of the 40 mg/L concentration for each individual animal was used as the selection trait. The lines diverged in the first generation of selection and continued to diverge with each subsequent generation of selection pressure.

MASENS line mice were administered saline on habituation days 1 and 2, and MA on days 3, 5, 6, 9 and 11. On each day, their locomotor behavior was quantified in automated activity monitors for 15 minutes immediately after injection. The change in activity level between days 3 (their initial response to MA) and 11 (their final response to MA) was the measure of degree of MA-induced sensitization (Day 11 minus Day 3) used as the selection trait.

The lines diverged in the first generation of selection and have continued to modestly diverge. This is very exciting because our previous estimates of heritability for this trait were low (h2 = ~0.12).

Progress toward measurement of genetically correlated responses to selection: MA drinking and sensitization

Second-litter MADR mice were tested for MA sensitization and MASENS mice were tested for MA drinking in each generation of selection. For the MADR lines, we have completed testing of the S1 - S3 generations. There were no significant differences between the lines in acute sensitivity to the locomotor stimulant effects of MA, or in the magnitude of MA-induced sensitization. This suggests that the genes mediating MA drinking do not have a pleiotropic effect on sensitivity to the locomotor effects of acute or repeated MA.

However, a different conclusion is reached when examining the MASENS lines for MA drinking. We have completed the testing of the S1 and S2 generations of the MASENS lines for MA consumption. S1 MAHSENS mice consumed more MA than MALSENS mice. However, in the S2 generation, this line difference was reversed, as the MALSENS line consumed more MA than the MAHSENS line. We predicted such a reversal in previous work (Kamens et al. (2005) Genes Brain Behav 4:110-125), where we suggested that extreme sensitivity to the psychomotor stimulant effects of MA may make MA aversive. This suggests some pleiotropy in the genetic influence on MA drinking and sensitization when sensitization is used as the selection trait. We will continue to examine this as both selections are completed.

MA metabolism

Clearance of MA was examined in MADR S2 mice after a single or five MA injections (1 mg/kg each). Injection dose and frequency mimicked those for our sensitization studies. There was no significant difference between the lines in elimination rate after acute MA. After repeated administration of MA, the MAHDR mice had significantly higher peak blood MA levels, compared to MALDR mice 15 min after administration, but clearance rate was comparable. MA metabolism will be examined again in S4 mice to determine if this difference persists or intensifies with selection. A similar study is being performed in MASENS mice.

MA conditioned place preference

MA-induced conditioned place preference (CPP) was examined in S3 MADR mice. This experiment tested the hypothesis that selection for differences in MA drinking resulted in changes in sensitivity to the reinforcing effects of MA. Tactile conditioning cues (i.e., floor textures) were used and mice were given 6 MA conditioning trials (one floor texture) interspersed with 6 saline conditioning trials (alternate floor texture), and then a final test (both floor textures present).

MAHDR mice exhibited significant preference for the MA-paired floor, an effect not seen in MALDR mice that exhibited no preference for either floor texture. These data are very exciting as they suggest that differences in MA drinking do index differences in sensitivity to the reinforcing effects of MA.

Cocaine drinking

Cocaine drinking was examined in S2 MADR mice. Procedures were exactly as performed for measurement of MA drinking. The lines did not differ significantly, but there was a trend in the expected direction of MAHDR mice consuming more cocaine, compared to MALDR mice. Tastant tests were also performed that examined consumption and preference for saccharin, quinine, and potassium chloride; no significant differences were found. This suggests that the differences in MA consumption are not due to differences in general taste sensitivity or preference. These studies are underway in the MASENS lines.

Cocaine sensitization

Cocaine sensitization was examined in S2 MADR mice (10 mg/kg). There was no significant difference between the lines in acute response to cocaine. Analyses across days revealed significant differences in activity between the lines after the 3rd-5th cocaine injections, with MALDR mice exhibiting a greater response to cocaine than the MAHDR mice. These results are consistent with our previously published work suggesting that high sensitivity to sensitization may lead to reduced drug consumption. S2 MASENS lines did not differ in response to acute or chronic cocaine. This will be examined in S4.

Ongoing work

MADR mice have been provided to Dr. Mitchell for impulsivity testing and Dr. Mark for i.c.v. MA self-administration (Component 5). Brain tissue has been delivered to Dr. Janowsky (Component 5) for examination of dopamine and ACh transporters and receptors. DNA will be collected from S4 mice of both lines in the next year for QTL mapping (Component 6). In addition, brains will be collected for laser dissection of specific brain regions for microarray gene expression analysis (Component 6) in the coming year.

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