GABABR/GABAB receptor

Lactate dehydrogenase (LDH) activity was significantly reduced in striatal cells (HD19 or HD43) expressing wild-type or heterogeneous huntingtin after being exposed to 1, 10 μM of baclofen for 24 hours. This suggests that the cells were more viable. In HD43 cells, baclofen dramatically boosts both cell survival and chymotrypsin-like pancreatic body activity [3].

In YAC128 HD buttons, baclofen (i.p. ; 10 μg/g; twice daily for three days) improves motor deficits [3].

Proteasome activity determination [3]
Proteasome activity was determined by measuring the fluorescence of 7-amido-4-methylcoumarin (AMC; excitation 380 nm, emission 460 nm) generated from peptide-AMC linked substrates. Reactions were conducted in a final volume of 200 μl of buffer including 50 mM Tris–HCl (pH 7.5) and 1 mM EDTA. After adding samples to the reaction mixtures, reactions were initiated by adding the substrate Suc-Leu-Leu-Val-Try-AMC (65 μM) to measure chymotrypsin-like activity. Reactions progressed for 360 min at 25 °C. Enzymatic activity was expressed as fluorescence units (FU)/mg/min of protein.

Cell culture and in vitro baclofen treatment [3]
Tet-off inducible wild type (HD19, 26 CAG repeats) and mutant (HD43, 105 CAG repeats) striatal cells were cultured at 33 °C in Dulbecco’s Modified Eagle’s Medium (DMEM; Hyclone, Logan, UT) supplemented with 10% fetal bovine serum (Hyclone), 2 mM l-glutamine (Sigma, St. Louis, MO), penicillin, and streptomycin. Expression of the huntingtin gene was induced by administration of doxycycline (1 μg/ml) for 24 h. The selective GABAB receptor agonist, baclofen (RS-baclofen; Tocris Cookson, Ellisville, MO), was administered to cultured cells at the indicated concentrations. After 24 h of administration, cell culture medium was collected for cell viability assays. Cells were harvested and lysed in a homogenization buffer (50 mM Tris (pH 8.0), 150 mM NaCl, 5 mM EDTA, 1% Triton X-100) containing the following protease and phosphatase inhibitors: 10 μg/ml aprotinin, 25 μg/ml leupeptin, 10 μg/ml pepstatin, 10 μg/ml phenylmethanesulfonyl fluoride, 50 mM sodium fluoride, 50 mM sodium orthovanadate) for protein sample preparation.
Cell viability determination[3]
We determined cell viability by performing a lactate dehydrogenase (LDH) assay on the collected cell medium, as per the manufacturer’s instructions (Roche, Mannheim, Germany). LDH activities of control or baclofen-treated striatal cells were measured by absorbance at 490 nm.

Animal/Disease Models: Wild type (WT) and mutant (MT) male YAC128 mice 13-18 months old [3]
Doses: 10 μg/g
Route of Administration: IP; twice (two times) daily at 9:00 AM and 5:00 PM , for 3 days; then a single dose was administered at 9:00 am on the fourth day.
Experimental Results: Motor deficits in YAC128 HD transgenic mice were improved. Increased proteasome activity and diminished neuronal intranuclear inclusions (NII) in YAC128 HD transgenic mice.

Absorption, Distribution and Excretion
Baclofen has an oral bioavailability of 70% to 85%. Following oral administration, it is rapidly absorbed through the gastrointestinal tract with peak plasma concentrations being reached two to three hours after ingestion. Peak effect is observed about four hours after intrathecal administration. The absorption is dose-dependent and increases with higher doses. There is intersubject variation in absorption. Administration of oral baclofen suspension with a high-fat meal resulted in 9% decrease in AUC and 33% decrease in Cmax compared to the fasted state.
About 70-80% of baclofen is eliminated in an unchanged form by renal excretion within 72 hours of administration. About 5% of the dose is excreted via the kidneys as metabolites. There is intersubject variation in elimination.
The volume of distribution of baclofen is 0.7 L/kg. As baclofen is mainly water-soluble, it does not readily cross the blood-brain barrier. Drug concentrations of baclofen in the cerebrospinal fluid are approximately 8.5 times lower than in the plasma.
The systemic clearance (CL/F) was 180 mL/min and the renal clearance was 103 mL/min following oral administration.

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Metabolism / Metabolites
Approximately 15% of the oral dose is metabolized in the liver, mainly by deamination. Deamination yields the main metabolite, β-(p-chlorophenyl)-4-hydroxybutyric acid, which is pharmacologically inactive.
~ 15% of the dose is metabolized in the liver, primarily by deamination. 70-80% of the dose is excreted unchanged or as metabolites in urine and the remainder is excreted in feces. Oral Baclofen is readily absorbed from the gastrointestinal tract. After oral administration, baclofen appears in the blodd within half an our. It is fairly distributed in most organs and body tissues. After oral administration of baclofen, about 85% is excreted unchanged in the urine and feces and the remainder is oxidatively dearninated in the liver to produce beta-(p-chlorophenyl)-gamma-hydroxybutyric acid as a major metabolite. (L1322).
Route of Elimination: In a study using radiolabeled baclofen, approximately 85% of the dose was excreted unchanged in the urine and feces.
Baclofen is excreted primarily by the kidney as unchanged drug; 70 - 80% of a dose appears in the urine as unchanged drug. The remainder is excreted as unchanged drug in the feces or as metabolites in the urine and feces.
Half Life: 2.5-4 hours

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Biological Half-Life
The half-life is 2-6 hours after oral administration and 1-5 hours following intrathecal administration. The apparent elimination half-life of baclofen oral suspension or granules is about 5.6 hours.

Toxicity Summary
Baclofen is a direct agonist at GABAB receptors. The precise mechanism of action of Baclofen is not fully known. It is capable of inhibiting both monosynaptic and polysynaptic reflexes at the spinal level, possibly by hyperpolarization of afferent terminals, although actions at supraspinal sites may also occur and contribute to its clinical effect.

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Toxicity Data
LD50: 45 mg/kg (Intravenous, Mouse) (A308)
LD50: 78 mg/kg (Intravenous, Rat) (A308)

[1]. A deeper insight into how GABA-B receptor agonism via baclofen may affect alcohol seeking and consumption: lessons learned from a human laboratory investigation. Mol Psychiatry. 2018 Oct 31.

[2]. Baclofen prevents MDMA-induced rise in core body temperature in rats. Drug Alcohol Depend, 2004. 74(1): p. 89-96.

[3]. Baclofen, a GABAB receptor agonist, enhances ubiquitin-proteasome system functioning and neuronal survival in Huntington's disease model mice. Biochem Biophys Res Commun. 2014 Jan 10;443(2):706-11.

Pharmacodynamics
Baclofen is an antispasmodic agent that induces muscle relaxation. It reduces the release of excitatory neurotransmitters in the pre-synaptic neurons and stimulates inhibitory neuronal signals in the post-synaptic neurons. Oral formulations of baclofen are the most commonly used form of the drug. In one cross-section study, intrathecal baclofen was more effective than oral baclofen in relieving spasticity directly at the level of the spinal cord. Baclofen has CNS depression properties and can cause sedation with tolerance, somnolence, ataxia, and respiratory and cardiovascular depression. Baclofen also mediates some antinociceptive effects and stimulates gastric acid secretion. Baclofen exhibits anti-inflammatory and neuroprotective activities: it inhibits the release of pro-inflammatory cytokines from microglia and astrocytes, and decreases oxidative stress in rats.

C10H12CLNO2

213.66

213.055

C, 56.21; H, 5.66; Cl, 16.59; N, 6.56; O, 14.98

1134-47-0

(R)-Baclofen;69308-37-8;Baclofen-d4;1189938-30-4;(R)-Baclofen hydrochloride;63701-55-3;Baclofen hydrochloride;28311-31-1

2284

White to off-white soild

1.3±0.1 g/cm3

364.3±32.0 °C at 760 mmHg

208-210°C

174.1±25.1 °C

0.0±0.9 mmHg at 25°C

1.577

1.56

2

3

4

14

191

0

O=C(O)CC(C1=CC=C(Cl)C=C1)CN

KPYSYYIEGFHWSV-UHFFFAOYSA-N

InChI=1S/C10H12ClNO2/c11-9-3-1-7(2-4-9)8(6-12)5-10(13)14/h1-4,8H,5-6,12H2,(H,13,14)

4-amino-3-(4-chlorophenyl)butanoic acid

Baclofen Lioresal, Liofen, Gablofen Baclon Kemstro

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

DMSO : ~4.81 mg/mL (~22.51 mM)
H2O : ~2 mg/mL (~9.36 mM)
0.1 M HCL: ~10 mg/mL (~46.8 mM)

Solubility in Formulation 1: 2.5 mg/mL (11.70 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication (<60°C).

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Solubility in Formulation 2: ~2.5 mg/mL (11.7 mM) in PBS

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