Journal:

Article Title: Muscarinic Acetylcholine Receptor Subtype Expression in Avian Vestibular Hair Cells, Nerve Terminals and Ganglion Cells

doi: 10.1016/j.neuroscience.2007.02.019

Figure Lengend Snippet: Antibodies and Antigens Used for Muscarinic Receptor Subtype Studies

Article Snippet: The primary antibodies included rabbit anti-M1-mAChR polyclonal antibody (Biodesign International, Cat. # Q4A173R, Saco, ME, USA), rabbit anti-M2-mAChR antibody (Alomone Labs Ltd., Cat. # AMR-002, Jerusalem, Israel), rabbit anti-M3-mAChR antibody (Alomone Labs Ltd., Cat. # AMR-006, Jerusalem, Israel), mouse anti-M4-mAChR monoclonal antibody (Chemicon International, Cat. # MAB1578, Temecular, CA) and rabbit anti-M5-mAChR antibody (Biodesign International, Cat. # Q4A871R, Saco, ME, USA).

Techniques:

Journal:

Article Title: Muscarinic Acetylcholine Receptor Subtype Expression in Avian Vestibular Hair Cells, Nerve Terminals and Ganglion Cells

doi: 10.1016/j.neuroscience.2007.02.019

Figure Lengend Snippet: Fluorescent labeling of mAChR subtype expression on vestibular peripheral structures in the crista, utricle and ganglion. The results shown here are the anti-M4 antibody reacted with a red fluorescent secondary antibody while other mAChR subtypes reacted with a green secondary antibody. No differences were found when the red/green fluorescent secondary antibodies were exchanged. The co-expression of M4 with M1, M3 and M5 on nerve fibers, nerve terminals and ganglion cells are marked by arrows. Myelin sheaths and Schwann cells surrounding ganglion cells are pointed at by single arrowheads. Cuticular plates w/ or w/o protruding cilia are pointed at by double arrows. The labeling above the row of hair cells in utricle (especially in E2) is the artifact induced by labeling the otoconial membrane which was not removed. Scale bars = 10 μm.

Article Snippet: The primary antibodies included rabbit anti-M1-mAChR polyclonal antibody (Biodesign International, Cat. # Q4A173R, Saco, ME, USA), rabbit anti-M2-mAChR antibody (Alomone Labs Ltd., Cat. # AMR-002, Jerusalem, Israel), rabbit anti-M3-mAChR antibody (Alomone Labs Ltd., Cat. # AMR-006, Jerusalem, Israel), mouse anti-M4-mAChR monoclonal antibody (Chemicon International, Cat. # MAB1578, Temecular, CA) and rabbit anti-M5-mAChR antibody (Biodesign International, Cat. # Q4A871R, Saco, ME, USA).

Techniques: Labeling, Expressing

Journal:

Article Title: Muscarinic Acetylcholine Receptor Subtype Expression in Avian Vestibular Hair Cells, Nerve Terminals and Ganglion Cells

doi: 10.1016/j.neuroscience.2007.02.019

Figure Lengend Snippet: DAB staining of (A) semicircular canal crista and (B) vestibular ganglion. The number in each panel corresponds to the mAChR subtype number. Myelin sheaths and Schwann cells circumscribe the nerve fibers and ganglion cells (black arrows). Nerve fibers expressing M1, M2, M4 and M5 traverse the basement membrane and terminate in the neuroepithelium (arrowheads). There is no M3 expression on peripheral nerve terminals inside of the neuroepithelium. The myelin sheath staining stops just beneath the basement membrane (double arrowheads in A-3). Scale bars = 10 μm.

Article Snippet: The primary antibodies included rabbit anti-M1-mAChR polyclonal antibody (Biodesign International, Cat. # Q4A173R, Saco, ME, USA), rabbit anti-M2-mAChR antibody (Alomone Labs Ltd., Cat. # AMR-002, Jerusalem, Israel), rabbit anti-M3-mAChR antibody (Alomone Labs Ltd., Cat. # AMR-006, Jerusalem, Israel), mouse anti-M4-mAChR monoclonal antibody (Chemicon International, Cat. # MAB1578, Temecular, CA) and rabbit anti-M5-mAChR antibody (Biodesign International, Cat. # Q4A871R, Saco, ME, USA).

Techniques: Staining, Expressing

Journal:

Article Title: Muscarinic Acetylcholine Receptor Subtype Expression in Avian Vestibular Hair Cells, Nerve Terminals and Ganglion Cells

doi: 10.1016/j.neuroscience.2007.02.019

Figure Lengend Snippet: Summary of mAChR subtype expression on the peripheral vestibular neurons

Article Snippet: The primary antibodies included rabbit anti-M1-mAChR polyclonal antibody (Biodesign International, Cat. # Q4A173R, Saco, ME, USA), rabbit anti-M2-mAChR antibody (Alomone Labs Ltd., Cat. # AMR-002, Jerusalem, Israel), rabbit anti-M3-mAChR antibody (Alomone Labs Ltd., Cat. # AMR-006, Jerusalem, Israel), mouse anti-M4-mAChR monoclonal antibody (Chemicon International, Cat. # MAB1578, Temecular, CA) and rabbit anti-M5-mAChR antibody (Biodesign International, Cat. # Q4A871R, Saco, ME, USA).

Techniques: Expressing

Journal: Brain and Behavior

Article Title: Most calbindin‐immunoreactive neurons, but few calretinin‐immunoreactive neurons, express the m1 acetylcholine receptor in the middle temporal visual area of the macaque monkey

doi: 10.1002/brb3.1071

Figure Lengend Snippet: Antibody details

Article Snippet: Anti‐m1 muscarinic acetylcholine receptor , GST fusion protein corresponding to aa227‐353 of human m1 ACh receptor (GSETPGKGGGSSSSSERSQPGAEGSPETPPGRCCRCCRAPRLLQAYSWKEEEEEDEGSMESLTSSEGEEPGESVVIKMPMVDPEAQAPTKQPPRSSPNTVKRPTKKGRDRAGKGQKPRGKEQLAKRK) , Alomone Labs (Jerusalem, Israel). Rabbit polyclonal. Catalog#AMR‐001 Lot#AN‐05 http://scicrunch.org/resolver/AB_2039993 , 1:1,000.

Techniques: Purification, Recombinant

Journal: Brain and Behavior

Article Title: Most calbindin‐immunoreactive neurons, but few calretinin‐immunoreactive neurons, express the m1 acetylcholine receptor in the middle temporal visual area of the macaque monkey

doi: 10.1002/brb3.1071

Figure Lengend Snippet: Quantification methods. The figure (a) shows m1 acetylcholine receptor immunoreactivity. Cells marked with a circle met counting criteria and were counted. The figure (b) shows immunoreactivity for calbindin‐D28k. Within this population are two subgroups: cells with darkly stained somata (marked with squares) and cells with faintly stained somata (marked with arrows). Only darkly stained cells were quantified. The merged image in (c) shows cells that were counted in both single‐label images, and thus are counted as dual‐labeled cells. Scale bar = 25 µm (all panels)

Article Snippet: Anti‐m1 muscarinic acetylcholine receptor , GST fusion protein corresponding to aa227‐353 of human m1 ACh receptor (GSETPGKGGGSSSSSERSQPGAEGSPETPPGRCCRCCRAPRLLQAYSWKEEEEEDEGSMESLTSSEGEEPGESVVIKMPMVDPEAQAPTKQPPRSSPNTVKRPTKKGRDRAGKGQKPRGKEQLAKRK) , Alomone Labs (Jerusalem, Israel). Rabbit polyclonal. Catalog#AMR‐001 Lot#AN‐05 http://scicrunch.org/resolver/AB_2039993 , 1:1,000.

Techniques: Staining, Labeling

Journal: Brain and Behavior

Article Title: Most calbindin‐immunoreactive neurons, but few calretinin‐immunoreactive neurons, express the m1 acetylcholine receptor in the middle temporal visual area of the macaque monkey

doi: 10.1002/brb3.1071

Figure Lengend Snippet: Laminar distributions of cells immunoreactive for the m1 acetylcholine receptor (a), calbindin‐D28k (b), and calretinin (c). Layer boundaries are marked to the left of each panel. m1‐immunoreactive neurons are more uniformly distributed throughout the tissue than are CB ‐ and CR ‐immunoreactive neurons, which are mostly expressed in layers II and III . Scale bar = 50 µm (all panels)

Article Snippet: Anti‐m1 muscarinic acetylcholine receptor , GST fusion protein corresponding to aa227‐353 of human m1 ACh receptor (GSETPGKGGGSSSSSERSQPGAEGSPETPPGRCCRCCRAPRLLQAYSWKEEEEEDEGSMESLTSSEGEEPGESVVIKMPMVDPEAQAPTKQPPRSSPNTVKRPTKKGRDRAGKGQKPRGKEQLAKRK) , Alomone Labs (Jerusalem, Israel). Rabbit polyclonal. Catalog#AMR‐001 Lot#AN‐05 http://scicrunch.org/resolver/AB_2039993 , 1:1,000.

Techniques:

Journal: Brain and Behavior

Article Title: Most calbindin‐immunoreactive neurons, but few calretinin‐immunoreactive neurons, express the m1 acetylcholine receptor in the middle temporal visual area of the macaque monkey

doi: 10.1002/brb3.1071

Figure Lengend Snippet: The figure (a) shows the percentage of the calbindin‐D28k ( CB ) and calretinin ( CR ) immunoreactive populations that are also immunoreactive (ir) for the m1 acetylcholine receptor (m1 AC hR), collapsed across all cortical layers. 55% of CB ‐ir neurons express m1 AC hRs, while 10% of CR ‐ir neurons express m1 AC hRs. The figure (b) shows the same data (52% of CB ‐ir neurons and 11% of CR ‐ir neurons), but for layers II and III only. The figure (c) shows the percentage of cells immunoreactive for m1 AC hRs that express CB or CR collapsed across all cortical layers. 2% of m1‐ir neurons express CB , while 0.5% of m1‐ir cells express CR . The figure (d) shows the same data (4% of m1 AC hR‐ir neurons express CB , while 1% express CR ), but for layers II and III only. Bars indicate standard error, and asterisks indicate statistical significance ( p < 0.01)

Article Snippet: Anti‐m1 muscarinic acetylcholine receptor , GST fusion protein corresponding to aa227‐353 of human m1 ACh receptor (GSETPGKGGGSSSSSERSQPGAEGSPETPPGRCCRCCRAPRLLQAYSWKEEEEEDEGSMESLTSSEGEEPGESVVIKMPMVDPEAQAPTKQPPRSSPNTVKRPTKKGRDRAGKGQKPRGKEQLAKRK) , Alomone Labs (Jerusalem, Israel). Rabbit polyclonal. Catalog#AMR‐001 Lot#AN‐05 http://scicrunch.org/resolver/AB_2039993 , 1:1,000.

Techniques:

Journal: Brain and Behavior

Article Title: Most calbindin‐immunoreactive neurons, but few calretinin‐immunoreactive neurons, express the m1 acetylcholine receptor in the middle temporal visual area of the macaque monkey

doi: 10.1002/brb3.1071

Figure Lengend Snippet: Neurons immunoreactive for the m1 acetylcholine receptor in layer III . Arrows mark cells that met counting criteria and were quantified. These neurons are characterized by strong somatic labeling appearing intense along the perimeter of the cell body. Scale bar = 20 µm

Article Snippet: Anti‐m1 muscarinic acetylcholine receptor , GST fusion protein corresponding to aa227‐353 of human m1 ACh receptor (GSETPGKGGGSSSSSERSQPGAEGSPETPPGRCCRCCRAPRLLQAYSWKEEEEEDEGSMESLTSSEGEEPGESVVIKMPMVDPEAQAPTKQPPRSSPNTVKRPTKKGRDRAGKGQKPRGKEQLAKRK) , Alomone Labs (Jerusalem, Israel). Rabbit polyclonal. Catalog#AMR‐001 Lot#AN‐05 http://scicrunch.org/resolver/AB_2039993 , 1:1,000.

Techniques: Labeling

Journal: Brain and Behavior

Article Title: Most calbindin‐immunoreactive neurons, but few calretinin‐immunoreactive neurons, express the m1 acetylcholine receptor in the middle temporal visual area of the macaque monkey

doi: 10.1002/brb3.1071

Figure Lengend Snippet: Comparison of the m1 acetylcholine receptor (m1 AC hR) expression by populations immunoreactive for calretinin ( CR ), calbindin‐D28k ( CB ), and parvalbumin ( PV ) in primary visual area V1 (dark gray) and middle temporal area MT (light gray). In V1, 40% of CR ‐immunoreactive neurons, 60% of CB ‐immunoreactive neurons, and 80% of PV ‐immunoreactive neurons express m1 AC hR. Those numbers in MT are 10%, 55%, and 75%, respectively

Article Snippet: Anti‐m1 muscarinic acetylcholine receptor , GST fusion protein corresponding to aa227‐353 of human m1 ACh receptor (GSETPGKGGGSSSSSERSQPGAEGSPETPPGRCCRCCRAPRLLQAYSWKEEEEEDEGSMESLTSSEGEEPGESVVIKMPMVDPEAQAPTKQPPRSSPNTVKRPTKKGRDRAGKGQKPRGKEQLAKRK) , Alomone Labs (Jerusalem, Israel). Rabbit polyclonal. Catalog#AMR‐001 Lot#AN‐05 http://scicrunch.org/resolver/AB_2039993 , 1:1,000.

Techniques: Expressing

Journal: bioRxiv

Article Title: Melanocortin 1 receptor regulates cholesterol and bile acid metabolism in the liver

doi: 10.1101/2022.11.08.515543

Figure Lengend Snippet: ( A) Immunostaining of MC1-R staining in the liver of chow-fed C57Bl/6J mouse. In control section, anti MC1-R antibody was replaced by purified normal rabbit IgG (isotype control). Scale bar, 50 μm. ( B) Quantitative real-time polymerase chain reaction (qPCR) analysis of Mc1r mRNA expression in the liver of chow- and Western diet-fed mice. ( C) Representative Western blots of MC1-R and β-actin (loading control) and quantification of MC1-R protein level in the liver of chow- and Western diet-fed mice. ( D) Schematic presentation of the loxP-flanked (floxed) Mc1r allele and the positions of forward and reverse primers used for PCR genotyping. PCR analysis of genomic DNA extracted from the liver of Alb-Cre-negative and -positive mice that were homozygous for the Mc1r floxed allele (Mc1r fl/fl ). The size of the recombined allele is ∼217 bp. ( E ) qPCR analysis of Mc1r expression in the liver of chow-fed Mc1r fl/fl , L-Mc1r +/- and L-Mc1r -/- mice at the age of 16 weeks. ( F, G ) Absolute liver weight and liver to body weight ratio (expressed as percentage of body weight) in chow-fed Mc1r fl/fl , L-Mc1r +/- and L-Mc1r -/- mice at the age of 16 weeks. Values are mean ± SEM, n = 5-10 mice per group in each graph. * P<0 . 05 and ** P<0 . 01 for the indicated comparisons by one-way ANOVA and Dunnet post hoc tests. L-Mc1r +/- indicates Alb-Cre +/- mice that were heterozygous for the Mc1r floxed allele; L-Mc1r -/- , hepatocyte-specific MC1-R knock-out mice.

Article Snippet: After blocking with Tris-Buffered Saline (Sigma-Aldrich) containing 0.1 % Tween ® 20 detergent (Sigma-Aldrich) and 5 % skimmed milk (Carl Roth) for 1 h at room temperature (RT), membranes were incubated with specific primary antibodies for MC1-R (Alomone Labs, #AMR-025), LDLR (NovusBio, Littleton, CO, USA, #NBP1-06709), SR-BI (NovusBio, #NB400-104), HMGCR (NovusBio, #NBP2-66888), DHCR7 (abcam, #ab103296), MRP4 (Cell Signaling Tech, Frankfurt, DE, #12857), StAR (Cell Signaling Tech, #8449), CYP8B1 (St John’s Laboratory Ltd, #STJ92607), phospho-AMPKα (Cell Signaling Tech, #2535), AMPKα (Cell Signaling Tech, #2532), phospho-Akt (R&D Systems, #AF887) and Akt (R&D Systems, #MAB2055) over-night at + 4°C.

Techniques: Immunostaining, Staining, Purification, Real-time Polymerase Chain Reaction, Expressing, Western Blot, Knock-Out

Journal: bioRxiv

Article Title: Melanocortin 1 receptor regulates cholesterol and bile acid metabolism in the liver

doi: 10.1101/2022.11.08.515543

Figure Lengend Snippet: ( A-C ) Representative Western blots and quantification of MC1-R protein level in HepG2 cells treated with palmitic acid (500 μM), LDL (200 μg/ml) or atorvastatin (10 μM) for 1, 3, 6 or 24 hours. (D) Quantification of free cholesterol content using filipin staining in HepG2 cells treated with α-MSH (1 μM) for 1, 3, 6 or 24 hours. (E) Quantification of free cholesterol content in HepG2 cells treated with different concentrations of α-MSH (0.1 nM, 10 nM or 1 μM) for 24 hours. (F, G) Quantification of LDL and HDL uptake in HepG2 cells treated with different concentrations (0.1 nM, 10 nM or 1 μM) of α-MSH for 24 hours. (H, I) qPCR analysis of LDLR and SCARB1 expression in HepG2 cells treated with different concentrations of α-MSH for 3, 6 or 24 hours. (J, K) Representative Western blots and quantification of LDL-R and SR-BI proteins levels in HepG2 cells treated with 1μM α-MSH for 1, 3, 6 or 24 hours. ( L ) Quantification of cell surface LDLR by flow cytometry in HepG2 cells treated with 1μM α-MSH for 24 hours. Values are mean ± SEM, * P<0 . 05 and ** P<0 . 01 for the indicated comparisons by one-way ANOVA and Dunnet post hoc tests ( A-K ) or by Student’s t test ( L ).

Article Snippet: After blocking with Tris-Buffered Saline (Sigma-Aldrich) containing 0.1 % Tween ® 20 detergent (Sigma-Aldrich) and 5 % skimmed milk (Carl Roth) for 1 h at room temperature (RT), membranes were incubated with specific primary antibodies for MC1-R (Alomone Labs, #AMR-025), LDLR (NovusBio, Littleton, CO, USA, #NBP1-06709), SR-BI (NovusBio, #NB400-104), HMGCR (NovusBio, #NBP2-66888), DHCR7 (abcam, #ab103296), MRP4 (Cell Signaling Tech, Frankfurt, DE, #12857), StAR (Cell Signaling Tech, #8449), CYP8B1 (St John’s Laboratory Ltd, #STJ92607), phospho-AMPKα (Cell Signaling Tech, #2535), AMPKα (Cell Signaling Tech, #2532), phospho-Akt (R&D Systems, #AF887) and Akt (R&D Systems, #MAB2055) over-night at + 4°C.

Techniques: Western Blot, Staining, Expressing, Flow Cytometry

Journal: bioRxiv

Article Title: Melanocortin 1 receptor regulates cholesterol and bile acid metabolism in the liver

doi: 10.1101/2022.11.08.515543

Figure Lengend Snippet: (A) Quantification of free cholesterol content using filipin staining in HepG2 cells treated with different concentrations of the selective MC1-R agonist LD211 (0.1 nM, 10 nM or 1 μM) for 24 hours. (B, C) Quantification of LDL and HDL uptake in HepG2 cells treated with different concentrations (0.1 nM, 10 nM or 1 μM) of LD211 for 24 hours. (D) qPCR analysis of LDLR and SCARB1 expression in HepG2 cells treated with 1 μM LD211 for 3, 6 or 24 hours. (E) Representative Western blots and quantification of LDL-R and SR-BI proteins levels in HepG2 cells treated with 1μM LD211 for 1, 3, 6 or 24 hours. ( F ) Quantification of cell surface LDLR by flow cytometry in HepG2 cells treated with 1μM LD211 for 24 hours. Values are mean ± SEM, * P<0 . 05 , ** P<0 . 01 , *** P<0 . 001 and **** P<0 . 0001 for the indicated comparisons by one-way ANOVA and Dunnet post hoc tests.

Article Snippet: After blocking with Tris-Buffered Saline (Sigma-Aldrich) containing 0.1 % Tween ® 20 detergent (Sigma-Aldrich) and 5 % skimmed milk (Carl Roth) for 1 h at room temperature (RT), membranes were incubated with specific primary antibodies for MC1-R (Alomone Labs, #AMR-025), LDLR (NovusBio, Littleton, CO, USA, #NBP1-06709), SR-BI (NovusBio, #NB400-104), HMGCR (NovusBio, #NBP2-66888), DHCR7 (abcam, #ab103296), MRP4 (Cell Signaling Tech, Frankfurt, DE, #12857), StAR (Cell Signaling Tech, #8449), CYP8B1 (St John’s Laboratory Ltd, #STJ92607), phospho-AMPKα (Cell Signaling Tech, #2535), AMPKα (Cell Signaling Tech, #2532), phospho-Akt (R&D Systems, #AF887) and Akt (R&D Systems, #MAB2055) over-night at + 4°C.

Techniques: Staining, Expressing, Western Blot, Flow Cytometry