• Modeling Mechanism：

  (RS)-(Tetrazol-5-yl)glycine (TZG), as a potent NMDA receptor agonist, induces pathology through multiple mechanisms: ① Overactivation of NMDA receptors in striatal neurons, leading to excitotoxicity, resulting in intracellular calcium overload (increased Ca²⁺ influx), and disrupting neuronal homeostasis; ② Induction of endothelial cell COX-2 expression, recruiting neutrophils to infiltrate the damaged area, with neutrophils expressing prostacyclin synthase (PGIS) and producing prostacyclin (PGI₂), forming a neuroprotective pathway of "endothelial COX-2-neutrophils-PGI₂"; ③ Induction of an inflammatory response in the striatal region (CD45⁺ leukocyte infiltration), while simultaneously leading to the formation of brain tissue lesions, mimicking the pathological features of excitatory brain injury associated with epileptic seizures.

• Related Products：

  (RS)-(Tetrazol-5-yl)glycine (T16801)

• Modeling Method：

  Experimental Subject：

  Mice: COX-2^(flox/flox), Tie2Cre COX-2^(flox/flox), LysMCre COX-2^(flox/flox) (conditional knockout mice) Age: 10–16 weeks Body weight: 25–30 g

  Dosage and Administration Route：

  ① Core modelling: (RS)-(Tetrazol-5-yl)glycine (5 μM), 10 μL/mouse, dissolved in PBS (pH 7.4), stereotaxic injection into right striatum (coordinates: 0.5 mm anterior to anterior fontanelle, +2.0 mm lateral, -4.0 mm ventral), single-dose administration;
  ② Surgical Procedure: Intraperitoneal anaesthesia with sodium pentobarbital (100 mg/kg), stereotaxic fixation of head, cranial drilling followed by microinjection at 0.8 μL/min using microinjection system, needle retention for 5 minutes to prevent backflow, wound suturing post-operatively;
  ③ Control treatment: Equal-volume PBS administered via identical injection protocol;
  ④ Intervention validation (optional):
  - PGI₂ agonist MRE-269 (1 mg/kg) administered intraperitoneally either 1/4 hour prior to modelling or post-modelling;
  - PGIS inhibitor fenpropirox (10 mg/kg) via intraperitoneal injection administered 1 hour prior to modelling.

  Dosing Frequency and Duration Model：

  Single-dose TZG injection

• Validation：

  Pathological markers: Tissue damage: HE staining showed obvious lesions in the striatum region, with the lesion volume peaking 24 hours after modeling. Intervention with phenylcyclopropane increased the lesion volume, while MRE-269 reduced it (P<0.05); Molecular markers: 12 hours after modeling, the level of stable metabolites of PGI₂ (6-ketoprostaglandin F1α) in the striatum of wild-type mice increased twofold (P<0.01), while the increase was not significant in Tie2Cre COX-2^flox/flox mice; CD45⁺ leukocytes and Ly6G⁺ neutrophils infiltrated more, and PGIS⁺ cells accumulated in the core of the lesion; Cellular markers: Calcium imaging showed that TZG induced a significant increase in calcium influx in striatal neurons (set as 100%), and MRE-269 pretreatment significantly reduced the amplitude and area under the curve of calcium influx (P<0.01); Specificity verification: Tie2Cre In COX-2^flox/flox mice (endothelial COX-2 knockout), TZG-induced PGI₂ production was reduced, PGIS⁺ cell infiltration was absent, and the lesion foci were enlarged, confirming the endothelial COX-2-dependent neuroprotective pathway.

*Precautions： The animals were euthanized at specific time points (4/8/12/24 hours) during the sampling process, and fresh brain tissue was handled on ice.

*References：An Y,et,al. Prostacyclin mediates endothelial COX-2-dependent neuroprotective effects during excitotoxic brain injury. J Inflamm Res. 2014 May 21;7:57-67.