(-)-Isoproterenol hydrochloride targets beta-1 (beta1) and beta-2 (beta2) adrenergic receptors with high affinity, and to a lesser extent beta-3 (beta3) receptors. It is a full agonist at these receptors, lacking significant activity at alpha-adrenergic receptors. Binding to beta1-adrenergic receptors (primarily in cardiac tissue) activates the Gs protein-cAMP-PKA signaling cascade, leading to increased heart rate (positive chronotropy), contractility (positive inotropy), and conduction velocity. Binding to beta2 receptors (in bronchial, vascular, and uterine smooth muscle) activates similar signaling pathways, resulting in smooth muscle relaxation (bronchodilation and vasodilation). The (-)-enantiomer is approximately 50-100 times more potent than the (+)-enantiomer at beta receptors, demonstrating stereoselectivity.

In cell-free radioligand binding assays, (-)-Isoproterenol hydrochloride competes with [3H]dihydroalprenolol or [¹2⁵I]cyanopindolol for binding to beta-adrenergic receptors in membrane preparations from various tissues (e.g., rat cardiac ventricle, lung). The Ki at beta1 receptors is approximately 50-100 nM, and at beta2 receptors is 5-50 nM, reflecting slightly higher potency at beta2. GTP (100 uM) reduces agonist binding affinity (converting high-affinity to low-affinity state), confirming G-protein coupling. In adenylate cyclase activity assays, isoproterenol (0.1 nM-100 uM) stimulates cAMP production in a concentration-dependent manner in cardiac or lung membrane preparations, with EC50 values of 10-100 nM. The effect is blocked by beta-antagonists such as propranolol (nonselective) or atenolol (beta1-selective) and butoxamine (beta2-selective), confirming receptor-mediated activity.

(-)-Isoproterenol hydrochloride (85 mg/kg; sc; daily for two consecutive days) can induce ventricular remodeling (VR) in rats[1].

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In cell-based assays using isolated rat or guinea pig cardiomyocytes, (-)-Isoproterenol hydrochloride (1 nM-10 uM) increases contractility (measured by edge-detection or sarcomere shortening) and intracellular cAMP levels (measured by ELISA or FRET-based biosensors). At higher concentrations (1-10 uM), it induces cardiomyocyte hypertrophy (increase in cell surface area, protein synthesis as measured by [3H]-leucine incorporation, and re-expression of fetal genes such as ANP and BNP) after 24-48 hours, serving as an in vitro model of cardiac hypertrophy. In bronchial smooth muscle cells or airway epithelial cells, isoproterenol (10 nM-10 uM) increases cAMP and relaxes pre-contracted cells (measured by cell impedance or traction force microscopy). In HEK293 cells transfected with beta2-adrenergic receptors, isoproterenol (10 nM-100 uM) induces receptor internalization (measured by flow cytometry or confocal microscopy of GFP-tagged receptors) and desensitization (reduced cAMP response upon re-challenge), providing a model for studying GPCR regulation. Cytotoxicity is minimal at concentrations below 10 uM over 24 hours.

For radioligand binding assays, beta-adrenergic receptors are prepared from rat cardiac ventricles (for beta1) or rat lung (for beta2). Tissues are homogenized in ice-cold buffer (25 mM Tris-HCl pH 7.4, 5 mM EDTA, 0.5 mM PMSF) and centrifuged at 40,000 × g for 20 minutes at 4degC. The pellet is washed twice and resuspended in binding buffer (50 mM Tris-HCl pH 7.4, 10 mM MgCl2, 0.1% ascorbate). Membranes (50-100 ug protein) are incubated with [3H]dihydroalprenolol (DHA, 2-5 nM) and varying concentrations of (-)-Isoproterenol hydrochloride (0.1 nM-1 mM) in a final volume of 250 uL for 60 minutes at 25degC. Non-specific binding is defined using 10 uM propranolol. Bound radioactivity is separated by rapid filtration through GF/B filters presoaked in 0.3% polyethyleneimine, washed three times with ice-cold buffer, and measured by liquid scintillation counting. Ki values are calculated using the Cheng-Prusoff equation. For adenylate cyclase assays, membranes (50 ug protein) are incubated in assay buffer (50 mM Tris-HCl pH 7.4, 5 mM MgCl2, 10 mM theophylline (PDE inhibitor), 1 mM ATP, 0.1 mM GTP, and an ATP-regenerating system) with isoproterenol (0.1 nM-100 uM) for 15 minutes at 30degC. The reaction is stopped by boiling for 3 minutes. cAMP generated is quantified by competitive ELISA. EC50 values are calculated by nonlinear regression.

For cell-based assays of cardiomyocyte contractility, neonatal rat ventricular myocytes (NRVMs) are isolated from 1-2 day old Sprague-Dawley rat pups. Hearts are excised, ventricles are minced and digested with trypsin and collagenase. Cells are pre-plated for 45 minutes to remove fibroblasts, then the enriched cardiomyocyte suspension is seeded in 35 mm dishes or 6-well plates coated with gelatin or laminin (2×10⁵ cells/cm2) and cultured in DMEM with 10% FBS, 1% penicillin/streptomycin, and 0.1 mM bromodeoxyuridine (to inhibit fibroblast proliferation) for 24-48 hours at 37degC with 5% CO2. Cells are serum-starved overnight before experiments. For contractility measurements, cells are placed in a temperature-controlled chamber (37degC) on an inverted microscope, and stimulated with a field stimulator (1 Hz, 5 ms pulses). After baseline recording, (-)-Isoproterenol hydrochloride (1-1000 nM) is added, and contractile amplitude (edge-detection software) and calcium transients (using Fura-2 or Fluo-4 loading) are recorded for 5-10 minutes. For hypertrophy studies, cells are treated with isoproterenol (1-10 uM) for 24-48 hours, then fixed and stained with rhodamine-phalloidin (to visualize F-actin) and DAPI (nuclei). Cell surface area is measured from microscopy images using ImageJ. For cAMP measurement, cells are treated with isoproterenol (0.1 nM-10 uM) for 10 minutes, then lysed, and cAMP is quantified by ELISA.

Animal/Disease Models: 180-200 g, 6-7 weeks, Healthy male Sprague Dawley rats[1]
Doses: 85 mg/kg
Route of Administration: S.c.; daily for two consecutive days
Experimental Results: Decreased the systolic blood pressure (SBP) levels, increased LV weight index (LVWI) and heart weight index (HWI), the average cross section area markedly increased, significant increased in the levels of NT-proBNP in the myocardium.

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A rat model of isoproterenol-induced cardiac hypertrophy and fibrosis is widely used to study heart failure. Male Sprague-Dawley rats (200-250g, n=10-15 per group) are injected subcutaneously with (-)-Isoproterenol hydrochloride (5 mg/kg/day dissolved in sterile saline) for 7-14 days. Control rats receive saline injections. Body weight is monitored daily. At the end of the treatment period, rats are anesthetized (e.g., with pentobarbital 50 mg/kg IP). Blood pressure and heart rate are measured via carotid artery cannulation. Electrocardiograms (ECG) are recorded to assess heart rate, QTc interval, and arrhythmias. Rats are euthanized, hearts are excised and weighed, and the heart weight-to-body weight ratio (HW/BW) is calculated as an index of cardiac hypertrophy. The left ventricle is dissected and fixed in 10% formalin for histological analysis: paraffin sections (5 um) are stained with H&E (for myocyte cross-sectional area), Masson's trichrome (for interstitial fibrosis, collagen volume fraction quantified by image analysis), and wheat germ agglutinin (WGA, for myocyte membrane and cross-sectional area). Gene expression of hypertrophic markers (ANP, BNP, beta-MHC) and fibrotic markers (collagen I, collagen III, TGF-beta) is measured by qPCR from LV tissue. Echocardiography (performed under light isoflurane anesthesia before sacrifice) measures left ventricular wall thickness, ejection fraction, and fractional shortening. Isoproterenol treatment causes significant increases in HW/BW (up to 30-50% increase), myocyte cross-sectional area, interstitial fibrosis, and reduced ejection fraction (30-50% decrease) compared to controls, modeling pathological cardiac remodeling. For mechanistic studies, rats are co-treated with beta-antagonists (propranolol 10 mg/kg/day IP, or atenolol 5 mg/kg/day) to confirm beta-receptor mediation.

Pharmacokinetic studies in humans and animals show that (-)-Isoproterenol hydrochloride is poorly bioavailable (F% <10%) when administered orally due to extensive first-pass metabolism in the gut and liver, primarily by catechol-O-methyltransferase (COMT). The compound is rapidly absorbed after parenteral administration: following intravenous injection in humans, the onset of action is immediate, with a short duration of 5-10 minutes due to rapid metabolism. The plasma half-life (t1/2) is 2-5 minutes. Volume of distribution (Vd) is approximately 0.5-1 L/kg. Plasma protein binding is low (20-40%). Isoproterenol is metabolized by COMT to 3-O-methylisoproterenol (an inactive metabolite) in the liver, gastrointestinal tract, and other tissues. It is also conjugated with sulfate. The metabolites are excreted in urine. In rodents, following IP administration (1-5 mg/kg), Tmax is 15-30 minutes, and plasma t1/2 is 10-20 minutes. For research use, (-)-Isoproterenol hydrochloride should be stored as a powder at -20degC, protected from light and moisture. Solutions should be prepared fresh in saline or 5% glucose (with 0.1% ascorbate as an antioxidant to prevent oxidation, which turns the solution pink/brown) and used within a few hours. Stock solutions in water (1-10 mM) can be stored at -80degC for up to 1 month.

(-)-Isoproterenol hydrochloride is toxic if ingested or injected at high doses. The oral LD50 in rats is approximately 3,000-5,000 mg/kg; IV LD50 is 30-50 mg/kg. The primary toxicities are cardiovascular: tachycardia, arrhythmias (premature ventricular contractions, ventricular tachycardia, atrial fibrillation), hypotension (due to beta2-mediated vasodilation), myocardial ischemia (due to increased oxygen demand), and cardiac hypertrophy with chronic use. At high doses (≥10 mg/kg in rats), isoproterenol causes myocardial necrosis (seen as focal areas of myocyte degeneration and calcification on histology), which is a standard toxicological endpoint in safety studies. It can also cause hyperglycemia (due to beta2-mediated hepatic glycogenolysis and insulin suppression) and hypokalemia (due to beta2-mediated potassium uptake into cells). The compound is classified as a hazardous substance (pharmaceutical ingredient). It is an irritant to skin and eyes (Category 2) and may cause respiratory irritation (Category 3) if inhaled. Isoproterenol has no known genotoxic or carcinogenic potential in standard assays (Ames test, micronucleus). For research handling, use appropriate PPE (nitrile gloves, lab coat, safety goggles), work in a fume hood to avoid inhalation of powder, and avoid skin contact as it may be absorbed. In case of accidental injection (needlestick), seek immediate medical attention. The compound is a controlled substance in some jurisdictions due to its cardiac effects and potential for misuse.

[1]. Cui XH, et al. Effect of Atractylodes macrocephala rhizoma on isoproterenol‑induced ventricular remodeling in rats. Mol Med Rep. 2018 Feb;17(2):2607-2613.

(-)-Isoproterenol hydrochloride (also known as (-)-Isoprenaline hydrochloride, isoproterenol HCl, Isuprel) is a pharmaceutical drug that has been approved in various countries for clinical use. It is indicated for: (1) bradycardia and heart block (temporary management, often as an IV infusion); (2) bronchospasm during anesthesia (as a bronchodilator); (3) cardiac stress testing (used to induce tachycardia to assess coronary artery function, though dobutamine is now more common); (4) torsade de pointes (as a temporary measure to increase heart rate). It is available as injectable solutions (0.2 mg/mL, 1 mg/5 mL) and as tablets (sublingual, though less common). The drug is not typically used as a first-line treatment due to its short half-life, arrhythmogenic potential, and the availability of more selective agents (e.g., dobutamine for inotropy, albuterol for bronchodilation). The (-)-enantiomer is the clinically used form. In research, isoproterenol is widely used as a tool to study beta-adrenergic signaling, cardiac physiology, GPCR desensitization, and as a positive control for inducing cardiac hypertrophy and fibrosis in animal models. It is also used to test beta-blocker efficacy. CAS: 5984-95-2, molecular formula C11H18ClNO3 (as HCl salt). The compound should be stored at 2-8degC (short-term) or -20degC (long-term) in a sealed, desiccated container, protected from light. It is hygroscopic, and solutions are prone to oxidation (forming colored degradation products). Purity is typically >98% by HPLC for research grades.

C11H18CLNO3

247.72

5984-95-2

White to off-white solid powder

441.5ºC at 760 mmHg

175ºC (dec.)(lit.)

220.8ºC

2.322

CC(C)NCC(C1=CC(=C(C=C1)O)O)O.Cl

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: (1). Please store this product in a sealed and protected environment, avoid exposure to moisture.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~125 mg/mL (~504.60 mM; with ultrasonication)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)