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5. GENETIC BASIS OF SUBSTANCE DEPENDENCE

application of the transgenic approach depends on the understanding of
mechanisms regulating gene expression in the mouse, which is currently
relatively limited (Quinn, 1996; Spergel et al., 2001).
The gene knockout methodology allows the deletion of a gene – or a
fragment of the gene – from the animal chromosomes. In this methodology
a mutated copy of the gene of interest is introduced into cultured mouse
embryonic stem cells. Through homologous recombination the mutated gene
becomes integrated into the genome of the stem cell, and disrupts (or
modifies, in the so-called knock-in technology) its function. Stem cells in
which the gene is disrupted are injected into blastocysts, which are then
implanted into foster mothers. Resulting mice have the gene disrupted in
some but not all cells, and are further bred to create knockout lines of mice
(Capecchi, 1994; Picciotto & Wickman, 1998).
Transgenic and knockout mice can serve as powerful research tools to
observe the effects of gene modifications. However, results from transgenic
and knockout studies need to be interpreted with caution for several reasons.
The site of integration for the transgene into the mouse chromosome is
random, and is as yet impossible to control. Therefore, some of the
phenotypes observed in transgenic mice can be due to the functions of the
transgene, but some can also be due to the disruption of the gene in which
the transgene has been integrated. Creation of several transgenic lines is
therefore necessary to verify that the observed phenotype is indeed due to
the transgene (Bowers, 2000). The knockout technology does not have the
problem of random transgene integration because the mutation in these mice
is targeted to a specific gene. However, it has other problems, for example,
the problem with background genotype (Crawley et al., 1997).
Both transgenic and knockout approaches are also faced with a problem
of developmental compensation. That is, while the gene that is modified or
over-expressed in the mutant animal could be important in the investigated
phenotype, compensatory mechanisms could also occur during development
(e.g. when a subunit of a receptor is knocked-out, another subunit could be
over-expressed and compensate for the absence of the knocked-out subunit).
If such compensation is occurring, the predicted change in the phenotype of
the mutant mice will not occur. New methodologies, including inducible
and brain region-specific transgenic and knockout approaches, are being
developed, and should in future alleviate many such problems (Sauer, 1998;
Le & Sauer, 2000).
Another approach used in animal studies is the quantitative trait loci (QTL)
analysis. Substance dependence is considered to be a quantitative trait in
which a combined action of multiple alleles leads to predisposition to
dependence. This approach does not assume any prior knowledge of genes
involved in substance-related disorders, and seeks to find them based on
related phenotypes. QTL analysis is analogous to linkage studies in humans.
As an example, inbred strains of mice that are genetically identical can be
crossed with other inbred strains, and the absence or presence of a mapped

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