1 The effects of the metabotropic glutamate receptor (mGluR) antagonist, L-2-amino-3-phosphonopropionate (L-AP3) on phosphoinositide turnover in neonatal rat cerebral cortex slices has been investigated. 2 accumulation which was similar in magnitude in both the absence and presence of IS, 3R-ACPD (300 UM). D-AP3 (1 mM) had no stimulatory effect alone and did not affect the response evoked by 1S, 3R-ACPD. L-AP3 (1 mM) also caused a large increase in Ins(1,4,5)P3 accumulation. The magnitude of the response (4-5 fold increase over basal) approached that evoked by a maximally effective concentration of 1S, 3R-ACPD, but differed substantially in the time-course of the response. The stimulatory effects of 1S, 3R-ACPD and L-AP3 on Ins(1,4,5)P3 accumulation were also similarly affected by decreases in extracellular calcium concentration. 4 Detailed analysis of the inositol phospholipid labelling pattern and the inositol (poly)phosphate isomeric species generated following addition of L-AP3 was also performed. In the continued presence of myo-[3H]-inositol, L-AP3 (1 mM) stimulated a significant increase in phosphatidylinositol labelling, but not that of the polyphosphoinositides, and the inositol (poly)phosphate profile suggested that substantial Ins(1,4,5)P3 metabolism occurs via both 5-phosphatase and 3-kinase routes. 5 A significant stimulatory effect of L-AP3 (1 mM) on [3H1-InsPx accumulation was also observed in neonatal rat hippocampus, and cerebral cortex and hippocampus slices prepared from adult rat brain. 6 These data demonstrate that whilst L-AP3 antagonizes mGluR-mediated phosphoinositide responses at concentrations of < 300 Mm, higher concentrations substantially stimulate this response. The ability of (±)-MCPG (1 mM) to attenuate significantly L-AP3-stimulated [3H]-InsPx accumulation, suggests that both the inhibitory and stimulatory effects of L-AP3 may be mediated by mGluRs.
It is suggested that mice exhibiting low physical activity levels overexpress Drd1 and this may be related to activity. This study aimed to evaluate if vivo‐morpholino (Gene Tools LLC) would knock down Drd1 expression and increase activity. Animal procedures were conducted in conformance with the FASEB Statement of Principles for the use of animals in research and education. Male C3H/HeJ (n=6 control; n=6 morpholino) mice (Jackson Labs) ran on wheels. After one week, the control group received a saline IV injection while the morpholino group received an IV vivo‐morpholino injection (11mg•kg−1) for three days. Four days following injection half of the cohort was sacrificed. Soleus and nucleus accumbens were harvested and analyzed on a western blot. The remaining mice were sacrificed and analyzed one week later. Four days post injection the morpholino group had a significant (p=0.03) knockdown of Drd1 in the soleus. There was no difference in Drd1 expression in the nucleus accumbens (p=0.33) nor was there a difference between the control and morpholino group twelve days post injection (p=0.53). There was no difference in wheel running between the control and morpholino group (p=0.42). The use of vivo‐morpholinos is effective in knocking down Drd1 in muscle but not in the nucleus accumbens potentially due to the blood brain barrier. Furthermore our data suggest that the vivo‐morpholinos gene knock‐down is transient.
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