Ontogeny of Norepinephrine Transporter Expression and Antidepressant-Like Response to Desipramine in Wild-Type and Serotonin Transporter Mutant Mice

Abstract

Depression is a major public health concern with symptoms that are often poorly controlled by treatment with common antidepressants. This problem is compounded in juveniles and adolescents, because therapeutic options are limited to selective serotonin reuptake inhibitors (SSRIs). Moreover, therapeutic benefits of SSRIs are often especially limited in certain subpopulations of depressed patients, including children and carriers of low-expressing serotonin transporter (SERT) gene variants. Tricyclic antidepressants (TCAs) offer an alternative to SSRIs; however, how age and SERT expression influence antidepressant response to TCAs is not understood. We investigated the relation between antidepressant-like response to the TCA desipramine using the tail suspension test and saturation binding of [³H]nisoxetine to the norepinephrine transporter (NET), the primary target of desipramine, in juvenile (21 days postnatal [P21]), adolescent (P28), and adult (P90) wild-type (SERT+/+) mice. To model carriers of low-expressing SERT gene variants, we used mice with reduced SERT expression (SERT+/-) or lacking SERT (SERT-/-). The potency and maximal antidepressant-like effect of desipramine was greater in P21 mice than in P90 mice and was SERT genotype independent. NET expression decreased with age in the locus coeruleus and increased with age in several terminal regions (e.g., the cornu ammonis CA1 and CA3 regions of the hippocampus). Binding affinity of [³H]nisoxetine did not vary as a function of age or SERT genotype. These data show age-dependent shifts for desipramine to produce antidepressant-like effects that correlate with NET expression in the locus coeruleus and suggest that drugs with NET-blocking activity may be an effective alternative to SSRIs in juveniles.

Graphical abstract

Fig. 1.

Influence of age and SERT genotype on the antidepressant-like effect of DMI. (A–C) Dose-dependent reductions in immobility time in the TST in P21, P28, and P90 SERT+/+ mice (A), SERT+/− mice (B), and SERT−/− mice (C). (D) Data from (A) expressed as a percentage of vehicle control. (E) Data from (B) expressed as a percentage of vehicle control. (F) Data from (C) expressed as a percentage of vehicle control. Data obtained in males and females are pooled, because a multifactor ANOVA (sex, genotype, DMI) showed no main effect or interaction of sex with other factors (P > 0.05), with one exception between sex and genotype (P < 0.01); however, multiple comparisons failed to show significant sex differences for each genotype (P > 0.05). Data are means ± S.E.M. Filled symbols represent data points that are significantly different from the SERT genotype– and age-matched vehicle control as determined by the Dunnett post hoc multiple comparisons test after a two-factor ANOVA. *P < 0.05 (significant difference from SERT genotype–matched P90); **P < 0.01 (significant difference from SERT genotype–matched P90); ^#P < 0.05 (significant difference from SERT genotype–matched P28 with the Tukey post hoc multiple comparisons test after a two-factor ANOVA); ^##P < 0.01 (significant difference from SERT genotype–matched P28 with the Tukey post hoc multiple comparisons test after a two-factor ANOVA). Sample sizes per data point were as follows: SERT+/+, n = 21–31 (9–14 males and 10–20 females, pooled); SERT+/−, n = 16–20 (8–10 males and 8–12 females, pooled); and SERT−/−, n = 18–23 (8–10 males and 9–14 females, pooled).

Fig. 2.

Specific [³H]nisoxetine binding to NET in hippocampal regions as a function of age and SERT genotype. Brain sections from P21, P28, and P90 SERT-deficient mice incubated with the NET-specific ligand [³H]nisoxetine. Nonspecific binding was defined by mazindol (2.5 mM). (A) Representative coronal sections at the level of plate 47 (Paxinos and Franklin, 1997) in SERT+/+, SERT+/−, and SERT−/− mice aged P21, P28, or P90. The boxed area in (A) is enlarged in (B), which shows representative thionine-stained brain sections labeled with hippocampal regions quantified, which include the CA1, CA2, and CA3 regions and the dentate gyrus (DG). (C) Example of saturation binding isotherms used to calculate B_max and K_d values. Curves include specific [³H]nisoxetine binding values for the CA3 of P21, P28, and P90 SERT+/+ mice. There was no main effect of sex on B_max or K_d values, so male and female data are pooled (P > 0.05). B_max values are summarized in Fig. 4. There were no significant differences in K_d among ages or between SERT+/+ and SERT+/− mice. Sample sizes of mice per group were as follows: SERT+/+, n = 5–9 (4 males and 3–5 females, pooled); SERT+/−, n = 6–10 (3–5 males and 2–4 females, pooled); and SERT−/−, n = 4–7 (2–4 males and 2–4 females, pooled). See Table 2 and Fig. 4 for a summary of data.

Fig. 3.

Specific [³H]nisoxetine binding to NET in the locus coeruleus as a function of age and SERT genotype. Brain sections from P21, P28, and P90 mice were incubated with increasing concentrations of [³H]nisoxetine. Nonspecific binding was defined by mazindol (2.5µM). (A) Representative coronal sections at the level of plate 76 (Paxinos and Franklin, 1997) in SERT+/+, SERT+/−, and SERT−/− mice aged P21, P28, or P90. The boxed area in (A) is enlarged in (B), which shows a representative thionine-stained brain sections including the locus coeruleus (LC). (C) Example of saturation binding isotherms used to calculate B_max and K_d values. Curves include specific [³H]nisoxetine binding values for the LC of P21, P28, and P90 SERT+/+ mice. B_max values for this region are summarized in Fig. 4. There was no main effect of sex on B_max or K_d values, so male and female data are pooled (P > 0.05). Sample sizes of mice per group were as follows: SERT+/+, n = 5–9 (4 males and 3–5 females, pooled); SERT+/−, n = 6–10 (3–5 males and 2–4 females, pooled); and SERT−/−, n = 4–7 (2–4 males and 2–4 females, pooled). See Table 2 and Fig. 4 for a summary of data.

Fig. 4.

B_max for [³H]nisoxetine binding to SERT in SERT+/+ and SERT+/− mice aged P21, P28, and P90. B_max values from P21, P28, and P90 SERT+/+, SERT+/−, and SERT−/− mice in the CA1 region (A), CA2 region (B), CA3 region (C), dentate gyrus (D), and locus coeruleus (E) were determined from one-site curve fits as described in the Materials and Methods. Data are means ± S.E.M. *P < 0.05 (significant difference from SERT genotype–matched P90, Dunnett post hoc multiple comparisons test after a two-factor ANOVA for age and SERT genotype). Data are means ± S.E.M. pooled from male and female mice. Sample sizes of mice per group were as follows: SERT+/+, n = 5–9 (4 males and 3–5 females, pooled); SERT+/−, n = 6–10 (3–5 males and 2–4 females, pooled); and SERT−/−, n = 4–7 (2–4 males and 2–4 females, pooled).

Fig. 5.

Relationship between E_max values for DMI to produce antidepressant-like effects in the TST and B_max values for specific [³H]nisoxetine binding in the locus coeruleus as a function of age and SERT genotype. The CA1 region (A), CA2 region (B), CA3 region (C), dentate gyrus (D), and locus coeruleus (E) are shown. Relationship between E_max and B_max did not vary by SERT genotype; thus, one line was used to fit data regardless of genotype. Data are taken from Fig. 1, D–F (per data point: SERT+/+, n = 21–31; SERT+/−, n = 16–20; and SERT−/−, n = 18–23), and Fig. 4 (per age group: SERT+/+, n = 5–9; SERT+/−, n = 6–10; and SERT−/−, n = 4–7). Data are means ± S.E.M. (male and female data are pooled).