A greater role for the norepinephrine transporter than the serotonin transporter in murine nociception

Abstract

Norepinephrine and serotonin involvement in nociceptive functions is supported by observations of analgesic effects of norepinephrine transporter (NET) and serotonin transporter (SERT) inhibitors such as amitriptyline. However, the relative contribution of NET and SERT to baseline nociception, as well as amitriptyline analgesia, is unclear. Amitriptyline and morphine analgesia in wild-type (WT) mice and littermates with gene knockout (KO) of SERT, NET or both transporters was conducted using the hotplate and tail-flick tests. Hypoalgesia was observed in NET KO mice, and to a lesser extent in SERT KO mice. The magnitude of this hypoalgesia in NET KO mice was so profound that it limited the assessment of drug-induced analgesia. Nonetheless, the necessary exclusion of these subjects because of profound baseline hypoalgesia strongly supports the role of norepinephrine and NET in basal nociceptive behavior while indicating a much smaller role for serotonin and SERT. To further clarify the role of NET and SERT in basal nociceptive sensitivity further experiments were conducted in SERT KO and NET KO mice across a range of temperatures. NET KO mice were again found to have pronounced thermal hypoalgesia compared to WT mice in both the hotplate and tail-flick tests, while only limited effects were observed in SERT KO mice. Furthermore, in the acetic acid writhing test of visceral nociception pronounced hypoalgesia was again found in NET KO mice, but no change in SERT KO mice. As some of these effects may have resulted from developmental consequences of NET KO, the effects of the selective NET blocker nisoxetine and the selective SERT blocker fluoxetine were also examined in WT mice: only nisoxetine produced analgesia in these mice. Collectively these data suggest that NET has a far greater role in determining baseline analgesia, and perhaps other analgesic effects, than SERT in mice.

Figure 1. Baseline analgesic responses in combined NET/SERT KO mice tested for morphine analgesia

Increased baseline analgesic responses (latency) was observed in NET KO, but not SERT KO, mice for supraspinal analgesia in the hot-plate test (A) and spinal analgesia in the tail-flick test (B).

Figure 2. Morphine-induced analgesia in combined NET/SERT KO mice

The data represent analgesic responses to morphine (0–20 mg/kg SC) in NET/SERT KO mice for supraspinal analgesia in the hot-plate test (A) and spinal analgesia in the tail-flick test (B). In both tests NET KO, but not SERT KO, appeared to increase sensitivity to morphine, although this effect did not reach significance in the hot-plate test and was reversed by SERT KO in the tail-flick test.

Figure 3. Baseline analgesic responses in combined NET/SERT KO mice tested for amitriptyline analgesia

Baseline analgesic responses (latency) in NET/SERT KO mice for supraspinal analgesia in the hot-plate test (A) and spinal analgesia in the tail-flick test (B). NET KO produced large increases in latencies in both tests, while SERT KO produced a small increase in latencies only in the hot-plate test.

Figure 4. Amitriptyline-induced analgesia in combined NET/SERT KO mice

The data represent analgesic responses to amitriptyline (0–40 mg/kg IP) in NET/SERT KO mice for supraspinal analgesia in the hot-plate test (A) and spinal analgesia in the tail-flick test (B). In the hot-plate test all genotypes except NET −/− SERT +/− and NET −/− SERT −/− had significant amitriptyline analgesia. NET KO dose-dependently enhanced hot-plate analgesia. In the tail-flick test all genotypes except NET−/− SERT −/− had significant amitriptyline analgesia. As discussed in the text because of the exclusion of NET subjects because of high baseline analgesia these results must be considered tentatively.

Figure 5. Thermal nociceptive sensitivity in NET KO mice

Reduced baseline nociceptive sensitivity observed in NET KO mice (+/+, +/− and −/−) for supraspinal analgesia in the hot-plate test (A), 47 ºC to 54 ºC, and spinal analgesia in the tail-flick test (B), 45 ºC to 52 ºC. Data represent response latencies (in seconds).

Figure 6. Thermal nociceptive sensitivity in SERT KO mice

No differences in baseline nociceptive sensitivity were observed in SERT KO (+/+, +/− and −/−) mice for supraspinal analgesia in the hot-plate test (A), 47 ºC to 54 ºC, or spinal analgesia in the tail-flick test (B), 45 ºC to 52 ºC. Data represent response latencies (in seconds).

Figure 7. Baseline visceral nociceptive responses in NET and SERT KO mice

Reduced visceral nociceptive responses were observed in (A) NET KO mice (+/+, +/− and −/−), but not (B) SERT KO mice(+/+, +/− and −/−) after intraperitoneal injection with acetic acid. Data represent the number of writhes (abdominal contractions) observed in the first 20 minutes after injection.

Figure 8. Analgesic effects of Nisoxetine but not Fluoxetine

Nisoxetine produced significant analgesia in the hot-plate (8A) and tail-flick (8B) tests, but fluoxetine did not produce significant analgesia in either the hot-plate (8C) or tail-flick (8D) tests.