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Increased Brain Neurotensin and NTSR2 Lead to Weak Nociception in NTSR3/Sortilin Knockout Mice

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ABSTRACT

The neuropeptide neurotensin (NT) elicits numerous pharmacological effects through three different receptors (NTSR1, NTSR2, and NTSR3 also called sortilin). Pharmacological approaches and generation of NTSR1 and NTSR2-deficient mice allowed to determine the NT-induced antipsychotic like behavior, the inhibitory of weak fear memory and the nociceptive signaling in a rat formalin tonic pain model to NTSR1. Conversely, the effects of NT on thermal and tonic nociceptions were mediated by NTSR2. However, the role of NTSR3/sortilin on the neurotensinergic system was not investigated. Here, by using C57Bl/6J mouse model in which the gene coding for NTSR3/sortilin has been inactivated, we observed a modification of the expression of both NTSR2 and NT itself. Quantitative PCR and protein expression using Western blot analyses and AlphaLisa™ technology resulted in the observation that brain NTSR2 as well as brain and blood NT were 2-fold increased in KO mice leading to a resistance of these mice to thermal and chemical pain. These data confirm that NTSR3/sortilin interacts with other NT receptors (i.e., NTSR2) and that its deletion modifies also the affinity of this receptor to NT.

No MeSH data available.


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Analgesic responses of WT and NTSR3/sortilin KO mice (A) Hot plate test, mice were placed on a plate at a temperature of 55°C. Bars represent mean ± SEM of paw licking and jumping latencies. Paw licking latency, ***p < 0.001, n = 20; jumping latency, *p < 0.05, n = 20. (B) Writhes were counted over a 15 min period after intraperitoneal injection of 0.5% acetic acid after intraperitoneal injection of either vehicle (NaCl) or 100 μl of 1 μM JT212. The number of indicated writhes is the mean ± SEM from groups of 10–12 mice. ***p < 0.001, *p < 0.05, n.s: non-significant.
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Figure 4: Analgesic responses of WT and NTSR3/sortilin KO mice (A) Hot plate test, mice were placed on a plate at a temperature of 55°C. Bars represent mean ± SEM of paw licking and jumping latencies. Paw licking latency, ***p < 0.001, n = 20; jumping latency, *p < 0.05, n = 20. (B) Writhes were counted over a 15 min period after intraperitoneal injection of 0.5% acetic acid after intraperitoneal injection of either vehicle (NaCl) or 100 μl of 1 μM JT212. The number of indicated writhes is the mean ± SEM from groups of 10–12 mice. ***p < 0.001, *p < 0.05, n.s: non-significant.

Mentions: Since we observed an increase of both NT and NTSR2 in NTSR3/sortilin KO mice and that NTSR2 is mainly involved in NT-induced analgesia (Dubuc et al., 1999), we wondered whether NTSR2 is still functional using acute pain tests including chemical (writhing test) and thermal (paw licking) nociceptive tests. When mice were placed on the hot plate, the latency for paw licking increased from 8.9 ± 0.85 s for WT mice to 17.3 ± 0.95 s for KO mice (p < 0.001) (Figure 4A). Similarly, the latency to jump was 30.9 ± 1.5 s for WT mice and increased to 39.4 ± 3.7 s for KO mice (p = 0.038) (Figure 4A), suggesting a resistance to pain for KO-NTSR3 mice. When WT mice were subjected to the writhing test, the number of writhes/15 min was 38.5 ± 5.2 (Figure 4B). In KO-NTSR3 mice, the number of writhes was 13.8 ± 3.6, a value significantly different to that obtained in WT mice (p < 0.001). Therefore, we tested the effect of IP injection of JT212 (100 μl of a 1 μM solution) on the pain writhing test and as expected, JT212 significantly decreased the number of writhes to 22.6 ± 3.7 in WT mice (One way ANOVA, p = 0.028) (Figure 4B). In KO mice, the injection of the peptide was without significant effect on the number of writhes (12.5 ± 2.4 (p = 0.99) (Figure 4B) indicating that no further analgesic action of JT212 was measurable when animals were already desensitized.


Increased Brain Neurotensin and NTSR2 Lead to Weak Nociception in NTSR3/Sortilin Knockout Mice
Analgesic responses of WT and NTSR3/sortilin KO mice (A) Hot plate test, mice were placed on a plate at a temperature of 55°C. Bars represent mean ± SEM of paw licking and jumping latencies. Paw licking latency, ***p < 0.001, n = 20; jumping latency, *p < 0.05, n = 20. (B) Writhes were counted over a 15 min period after intraperitoneal injection of 0.5% acetic acid after intraperitoneal injection of either vehicle (NaCl) or 100 μl of 1 μM JT212. The number of indicated writhes is the mean ± SEM from groups of 10–12 mice. ***p < 0.001, *p < 0.05, n.s: non-significant.
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Related In: Results  -  Collection

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Figure 4: Analgesic responses of WT and NTSR3/sortilin KO mice (A) Hot plate test, mice were placed on a plate at a temperature of 55°C. Bars represent mean ± SEM of paw licking and jumping latencies. Paw licking latency, ***p < 0.001, n = 20; jumping latency, *p < 0.05, n = 20. (B) Writhes were counted over a 15 min period after intraperitoneal injection of 0.5% acetic acid after intraperitoneal injection of either vehicle (NaCl) or 100 μl of 1 μM JT212. The number of indicated writhes is the mean ± SEM from groups of 10–12 mice. ***p < 0.001, *p < 0.05, n.s: non-significant.
Mentions: Since we observed an increase of both NT and NTSR2 in NTSR3/sortilin KO mice and that NTSR2 is mainly involved in NT-induced analgesia (Dubuc et al., 1999), we wondered whether NTSR2 is still functional using acute pain tests including chemical (writhing test) and thermal (paw licking) nociceptive tests. When mice were placed on the hot plate, the latency for paw licking increased from 8.9 ± 0.85 s for WT mice to 17.3 ± 0.95 s for KO mice (p < 0.001) (Figure 4A). Similarly, the latency to jump was 30.9 ± 1.5 s for WT mice and increased to 39.4 ± 3.7 s for KO mice (p = 0.038) (Figure 4A), suggesting a resistance to pain for KO-NTSR3 mice. When WT mice were subjected to the writhing test, the number of writhes/15 min was 38.5 ± 5.2 (Figure 4B). In KO-NTSR3 mice, the number of writhes was 13.8 ± 3.6, a value significantly different to that obtained in WT mice (p < 0.001). Therefore, we tested the effect of IP injection of JT212 (100 μl of a 1 μM solution) on the pain writhing test and as expected, JT212 significantly decreased the number of writhes to 22.6 ± 3.7 in WT mice (One way ANOVA, p = 0.028) (Figure 4B). In KO mice, the injection of the peptide was without significant effect on the number of writhes (12.5 ± 2.4 (p = 0.99) (Figure 4B) indicating that no further analgesic action of JT212 was measurable when animals were already desensitized.

View Article: PubMed Central - PubMed

ABSTRACT

The neuropeptide neurotensin (NT) elicits numerous pharmacological effects through three different receptors (NTSR1, NTSR2, and NTSR3 also called sortilin). Pharmacological approaches and generation of NTSR1 and NTSR2-deficient mice allowed to determine the NT-induced antipsychotic like behavior, the inhibitory of weak fear memory and the nociceptive signaling in a rat formalin tonic pain model to NTSR1. Conversely, the effects of NT on thermal and tonic nociceptions were mediated by NTSR2. However, the role of NTSR3/sortilin on the neurotensinergic system was not investigated. Here, by using C57Bl/6J mouse model in which the gene coding for NTSR3/sortilin has been inactivated, we observed a modification of the expression of both NTSR2 and NT itself. Quantitative PCR and protein expression using Western blot analyses and AlphaLisa&trade; technology resulted in the observation that brain NTSR2 as well as brain and blood NT were 2-fold increased in KO mice leading to a resistance of these mice to thermal and chemical pain. These data confirm that NTSR3/sortilin interacts with other NT receptors (i.e., NTSR2) and that its deletion modifies also the affinity of this receptor to NT.

No MeSH data available.


Related in: MedlinePlus