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| Targets |
The primary target of the PACAP-38 (31-38) fragment is the PAC1 receptor (also known as ADCYAP1R1), a class B GPCR that preferentially binds PACAP over vasoactive intestinal peptide (VIP). This fragment binds to the PAC1 receptor with low micromolar affinity, acting as a partial agonist. Upon binding, it activates adenylate cyclase, increasing intracellular cAMP and elevating cytosolic Ca2+. This signaling pathway leads to increased phosphorylation of extracellular regulated kinase (ERK) and activation of the epidermal growth factor receptor (EGFR). It also increases alpha-secretase activity.
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| ln Vitro |
PACAP-38(31–38), human, mouse, and rat TFA (100 nM; 2 min; NCI-H838 cells) tyrosine phosphorylates EGFR, HER2, and ERK[1]. Inducing EGFR tyrosine phosphorylation with PACAP-38(31–38), human, mouse, and rat TFA (100 nM; 30 min; NCI-H838 cells) raises ROS levels by 51%[1]. NCI-H838 cell growth is stimulated by TFA (10 nM; 48 h) in human, mouse, and rat PACAP-38(31–38)[1]. Human, mouse, and rat TFA (300 nM; 4 h) together with PACAP-38(31–38) increases the production of APPsα in brain cells[2]. Through cAMP accumulation and an increase in cytosolic free calcium, PACAP-38(31–38), human, mouse, and rat TFA(0.01–10 nM; HEK 293 cells) stimulates neural cells express endogenous PAC1 receptors. It also induces elevation of the intracellular Ca2+ concentration in a dose-dependent manner with an EC50 value of 0.81 nM[2]. PACAP-38(31–38), human, mouse, and rat TFA (0.01 nM; 48 h) stimulates NPY release from SCG neuronal cells in a strong and effective manner[3]. Human, mouse, and rat TFA (100 nM; 14 d) along with PACAP-38(31–38) generates prolonged activated NPY and catecholamine secretionmpathetic neurons[3].
In primary sympathetic neuronal cell cultures from the superior cervical ganglion (SCG), PACAP-38 (31-38) TFA (0.01 nM; 48 hours) potently and effectively stimulates neuropeptide Y (NPY) release. The fragment (100 nM; 14 days) also generates prolonged and sustained NPY and catecholamine secretion from sympathetic neurons. These stimulatory effects are potent, efficacious, and sustained. The C-terminal sequence (31-38) appears to be important for the long-term neurotrophic effects of PACAP, particularly in promoting neuronal differentiation and survival. |
| ln Vivo |
The fragment is primarily used in ex vivo culture systems rather than whole-animal studies. In NGF-overexpressing (NGF-OE) transgenic mice, the full-length PACAP antagonist (6-38) has well-characterized effects, but the (31-38) fragment (since it is an agonist fragment, not the antagonist) is typically studied in isolated tissues. Administration of the fragment to sympathetic neurons in vivo stimulates the sustained production of NPY and catecholamines, promoting sympathetic neuron development, differentiation, and enhancing nerve growth factor (NGF) signaling.
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| Enzyme Assay |
This is generally not a standard protocol due to the fragment nature; the typical full-length receptor binding assay is described: Membranes from CHO or HEK293 cells stably expressing the human PAC1 receptor (5-20 ug protein) are incubated with 50-100 pM ¹2⁵I-PACAP-27 (radiolabeled ligand) in binding buffer (20 mM HEPES, pH 7.4, 100 mM NaCl, 5 mM MgCl2, 1 mM CaCl2, 0.1% BSA, 0.1 mg/mL bacitracin) for 60 min at room temperature in the presence of increasing concentrations (0.1 nM - 10 uM) of PACAP-38 (31-38) TFA or unlabeled PACAP-38. Bound radioligand is separated by rapid filtration through GF/C filters pre-soaked in 0.3% polyethylenimine. Filters are washed 3 times with ice-cold binding buffer. Radioactivity is quantified by gamma counter. Nonspecific binding is determined using 1 uM unlabeled PACAP-38. IC50 values are calculated by curve-fitting; Ki is calculated using the Cheng-Prusoff equation.
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| Cell Assay |
Cell Viability Assay[1]
Cell Types: NCI-H838 cells Tested Concentrations: 10 nM Incubation Duration: 48 hrs (hours) Experimental Results: Increased the number of NCI-H838 cells by 72%. Western Blot Analysis[1] Cell Types: NCI-H838 cells Tested Concentrations: 100 nM Incubation Duration: 2 min Experimental Results: Increased tyrosine phosphorylation of the EGFR, HER2, and ERK by 377, 299 and 216%, respectively. Primary sympathetic neurons are dissected from the superior cervical ganglia (SCG) of neonatal (P0-P2) rats or mice. SCG are dissociated by enzymatic digestion (collagenase/dispase, 30 min, 37degC) and trituration. Cells (2×10⁵ cells/well in 24-well plates) are cultured in Neurobasal medium with B27 supplement, 2 mM glutamine, and 50 ng/mL NGF. After 7 days in vitro (DIV7), neurons are washed and treated with PACAP-38 (31-38) TFA (0.01, 0.1, 1, 10, 100 nM) for 48 hours or 14 days (for long-term secretion studies). NPY and catecholamine levels in the culture medium are measured by ELISA. For calcium imaging, cells are loaded with Fura-2 AM (5 uM, 30 min, 37degC) and the ratio of fluorescence at 340/380 nm is recorded. A PAC1 antagonist (PACAP(6-38), 1 uM) should be co-incubated to confirm receptor specificity. |
| Animal Protocol |
Standard animal protocols exist for the full-length PACAP peptide, but the (31-38) fragment is typically used in ex vivo tissue culture. For sympathetic neuron stimulation in vivo, NGF-OE transgenic mice (which overexpress NGF) are used. PACAP-38 (31-38) TFA (100 nM in 10 uL PBS) is injected intrathecally (i.t.) or intraperitoneally (i.p.) daily for 14 days. The expression of NPY and tyrosine hydroxylase (TH) in sympathetic ganglia (superior cervical, stellate, celiac) is assessed by qPCR, Western blot, and immunohistochemistry. Plasma catecholamine and NPY levels are measured by ELISA. Sympathetic nerve density in target tissues (e.g., heart, vasculature) is evaluated by immunohistochemistry for TH and NPY.
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| ADME/Pharmacokinetics |
Pharmacokinetics of this short, linear 8-amino acid peptide are not reported. Its half-life in plasma or CSF is expected to be less than 10 minutes due to rapid degradation by peptidases. For prolonged effects (e.g., 14-day NPY secretion), the peptide must be administered daily or via continuous infusion (osmotic minipump). The TFA salt form ensures solubility in aqueous buffers. No detailed PK profiles are available; for in vivo use, the compound is typically used in ex vivo tissue culture models rather than whole-animal systemic pharmacology.
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| Toxicity/Toxicokinetics |
Specific toxicity data for this PACAP fragment are not available. At the concentrations used in cell culture (0.01-100 nM), no cytotoxicity or morphological changes are observed. Given that the compound is derived from an endogenous neuropeptide, toxicity is expected to be low. High concentrations may cause overstimulation of sympathetic neurons, leading to excessive NPY release and potential cardiovascular effects, but this has not been systematically studied.
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| References |
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| Additional Infomation |
PACAP-38 (31-38) TFA is strictly a research-grade peptide tool. The full-length PACAP-38 peptide is a neurotrophic factor and neurotransmitter with known roles in development, stress response, and neuroprotection. This C-terminal fragment serves as a valuable probe for mapping the functional domains of PACAP, especially the regions responsible for sustained versus transient cellular responses. The TFA salt is the standard form for peptide synthesis. The peptide is available in human, mouse, and rat sequences, which are identical in this region. Store at -20degC as lyophilized powder; reconstitute in sterile water or PBS for use.
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| Molecular Formula |
C49H84F3N17O13
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| Molecular Weight |
1176.29
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| Related CAS # |
PACAP-38 (31-38), human, mouse, rat;138764-85-9
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| Appearance |
Solid powder
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: Please store this product in a sealed and protected environment, avoid exposure to moisture. |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
H2O :~100 mg/mL (~85.01 mM)
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| Solubility (In Vivo) |
Solubility in Formulation 1: 100 mg/mL (85.01 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.
(Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 0.8501 mL | 4.2507 mL | 8.5013 mL | |
| 5 mM | 0.1700 mL | 0.8501 mL | 1.7003 mL | |
| 10 mM | 0.0850 mL | 0.4251 mL | 0.8501 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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