Figure 1.
Molecular structures Triple reuptake inhibitors.
Figure 2.
Synthesis of D-473.
Table 1.
Inhibition of Drugs at hDAT, hSERT, and hNET in HEK293 Cells as Determined in Uptake Assays.
Table 2.
Binding affinity of D-473 for CNS receptors.
Figure 3.
Inhibition of [3H]monoamine uptake by D-473 and D-142 in cells heterologously expressing hDAT, hSERT, or hNET (▪-, D-473; Δ-, D-142).
A) Monoamine uptake by DAT, B) Monoamine uptake by SERT, and C) Monoamine uptake by NET was assessed as described in Methods and plotted according to the Logistics equation in the Origin fitting software (see Reith et al., 2012). Points shown are those obtained in a representative experiment, performed in triplicate, which was replicated 8–16 times.
Figure 4.
Effect of sub-chronic oral administration of vehicle and D-473 on the duration of immobility in the forced swimming test in rats.
One way ANOVA analysis demonstrates significant effect among treatments: F (4, 24) = 10.33 (P<0.0001). Dunnett’s analysis showed that the effect of D-473 at three doses (10, 25 and 50 mg/kg) on immobility was statistically significant different compared to vehicle (P<0.01).
Figure 5.
Effects of D-473 (PO) and GBR 12909 (i.p), on locomotor activity (horizontal activity, HACTV, measured by the number of infrared photobeam interuptions).
D-473 was administered orally to the rats whereas Vehicle and GBR 12909 were injected i.p. The cumulative locomotor activities up to 90 min were measured. One way ANOVA analysis and subsequent Dunnett’s multiple comparisons testing demonstrates no significant difference between control and D-473 but a significant effect between control and GBR 12909: F (2,6) = 18.22 (P<0.0028). Asterisks indicate a statistically significant difference compared with Vehicle, **P<0.002. Each treatment group contains four to five rats.
Figure 6.
a) Average plasma concentration of D-473 and D-142 versus time following oral administration in male Sprague-Dawley rats at 25 mg/kg from a 3% HPßCD in water formulation. b) Time dependent concentration of D-473 in the brain following oral administration in male Sprague-Dawley rats at 25 mg/kg from a 3% HPßCD in water formulation.
Figure 7.
Time dependent effect of administration of D-473 (10 mg/kg, i.p) at time 0 (shown by arrow) on extracellular level of DA (Ο), 5-HT (▪), and NE (Δ); a) in rats Prefrontal cortex; and b) dorso lateral striatum.
Results are expressed as percent baseline with baseline values all averaging 100%. Each point represents mean ± standard error (SE) of the percentage of baseline from five rats. Statistical analysis was performed by t-test analysis of every point relative to baseline values (i.e. 100%) using Prism 6 (Graphpad Software Inc, La Jolla, CA). * (DA) p<0.03-0.009; œ (NE) p<0.05- 0.01; # (5HT) p<0.05- 0.0001. P values <0.05 were considered to be statistically significant.
Figure 8.
Time dependent effect of administration of D-473 (10 mg/kg, i.p) at time 0 (shown by arrow) on extracellular level of DOPAC (Ο), HVA (▪), and 5HIAA (Δ); a) in rats Prefrontal cortex; and b) Dorso lateral striatum.
Results are expressed as percent baseline with baseline values all averaging 100%. Each point represents mean ± standard error (SE) of the percentage of baseline from five rats. Statistical analysis was performed by t-test analysis of every point relative to baseline values (i.e. 100%) using Prism 6 (Graphpad Software Inc, La Jolla, CA). * (DOPAC) p<0.02-0.0004. # (HVA) p<0.02–0.001. P values <0.05 were considered to be statistically significant.