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Updated: May 31, 2026
Quantifying the Heterogeneous Distribution of a Synaptic Protein in the Mouse Brain Using Immunofluorescence
Published on: January 29, 2019
Vesicular monoamine transporter-1 (VMAT-1) mRNA and immunoreactive proteins in mouse brain
Karen M Ashe1, Wan-Ling Chiu, Ahmed M Khalifa
1Department of Biology, Virginia Commonwealth University, Richmond, VA 23284, USA.
Objective:
Vesicular monoamine transporter 1 (VMAT-1) mRNA and protein were examined (1) to determine whether adult mouse brain expresses full-length VMAT-1 mRNA that can be translated to functional transporter protein and (2) to compare immunoreactive VMAT-1 proteins in brain and adrenal.
Methods:
VMAT-1 mRNA was detected in mouse brain with RT-PCR. The cDNA was sequenced, cloned into an expression vector, transfected into COS-1 cells, and cell protein was assayed for VMAT-1 activity. Immunoreactive proteins were examined on western blots probed with four different antibodies to VMAT-1.
Results:
Sequencing confirmed identity of the entire coding sequences of VMAT-1 cDNA from mouse medulla oblongata/pons and adrenal to a Gen-Bank reference sequence. Transfection of the brain cDNA into COS-1 cells resulted in transporter activity that was blocked by the VMAT inhibitor reserpine and a proton ionophore, but not by tetrabenazine, which has a high affinity for VMAT-2. Antibodies to either the C- or N- terminus of VMAT-1 detected two proteins (73 and 55 kD) in transfected COS-1 cells. The C-terminal antibodies detected both proteins in extracts of mouse medulla/pons, cortex, hypothalamus, and cerebellum but only the 73 kD protein and higher molecular weight immunoreactive proteins in mouse adrenal and rat PC12 cells, which are positive controls for rodent VMAT-1.
Conclusions:
These findings demonstrate that a functional VMAT-1 mRNA coding sequence is expressed in mouse brain and suggest processing of VMAT-1 protein differs in mouse adrenal and brain.
Insights
Functional vesicular monoamine transporter 1 (VMAT-1) mRNA is expressed in the adult mouse brain, leading to active transporter proteins. However, VMAT-1 protein processing appears to differ between the mouse brain and adrenal gland.
Area of Science:
- Neuroscience
- Molecular Biology
- Biochemistry
Background:
- Vesicular monoamine transporter 1 (VMAT-1) plays a crucial role in neurotransmitter packaging.
- Understanding VMAT-1 expression and function in the adult brain is essential for neurological research.
Purpose of the Study:
- To determine if adult mouse brain expresses functional VMAT-1 mRNA and protein.
- To compare VMAT-1 protein expression in the brain versus the adrenal gland.
Main Methods:
- RT-PCR was used to detect VMAT-1 mRNA in mouse brain.
- cDNA sequencing, expression vector cloning, and COS-1 cell transfection were performed.
- Western blotting with specific antibodies was used to analyze VMAT-1 proteins.
Main Results:
- Sequencing confirmed the full coding sequence of VMAT-1 cDNA in mouse brain and adrenal glands.
- Transfected brain cDNA yielded functional VMAT-1 activity, inhibited by reserpine but not tetrabenazine.
- Two VMAT-1 proteins (73 and 55 kD) were detected in brain extracts, while the adrenal gland showed predominantly the 73 kD form.
Conclusions:
- Functional VMAT-1 mRNA is expressed in the adult mouse brain.
- Evidence suggests differential processing of VMAT-1 protein in the mouse brain compared to the adrenal gland.
