Protocol for live neuron imaging analysis of basal surface fraction and dynamic availability of the dopamine transporter using DAT-pHluorin

Elnaz Khezerlou; Jacqueline Saenz; Sanjana Surya Prakash; Ping Yue Pan

doi:10.1016/j.xpro.2024.103358

Protocol for live neuron imaging analysis of basal surface fraction and dynamic availability of the dopamine transporter using DAT-pHluorin

Elnaz Khezerlou
, Jacqueline Saenz
, Sanjana Surya Prakash
, Ping Yue Pan

Research output: Contribution to journal › Article › peer-review

Abstract

Surface availability of the dopamine (DA) transporter (DAT) critically influences DA transmission. Here, we present a protocol that describes the preparation of mouse ventral midbrain neurons, the expression of a new optical sensor, DAT-pHluorin, and the utilization of this sensor to analyze the surface availability of DAT in live neurons via fluorescent microscopy. This approach allows quantitative measures of basal surface DAT fraction under genetic backgrounds of interest and live trafficking of DAT in response to psychoactive substances. For complete details on the use and execution of this protocol, please refer to Saenz et al.¹

Original language	English
Article number	103358
Journal	STAR Protocols
Volume	5
Issue number	4
DOIs	https://doi.org/10.1016/j.xpro.2024.103358
State	Published - 20 Dec 2024
Externally published	Yes

Keywords

cell biology
cell culture
cell-based assays

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10.1016/j.xpro.2024.103358

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title = "Protocol for live neuron imaging analysis of basal surface fraction and dynamic availability of the dopamine transporter using DAT-pHluorin",

abstract = "Surface availability of the dopamine (DA) transporter (DAT) critically influences DA transmission. Here, we present a protocol that describes the preparation of mouse ventral midbrain neurons, the expression of a new optical sensor, DAT-pHluorin, and the utilization of this sensor to analyze the surface availability of DAT in live neurons via fluorescent microscopy. This approach allows quantitative measures of basal surface DAT fraction under genetic backgrounds of interest and live trafficking of DAT in response to psychoactive substances. For complete details on the use and execution of this protocol, please refer to Saenz et al.1",

keywords = "cell biology, cell culture, cell-based assays",

author = "Elnaz Khezerlou and Jacqueline Saenz and Prakash, \{Sanjana Surya\} and Pan, \{Ping Yue\}",

note = "Publisher Copyright: {\textcopyright} 2024 Rutgers Robert Wood Johnson Medical School",

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AU - Khezerlou, Elnaz

AU - Saenz, Jacqueline

AU - Prakash, Sanjana Surya

AU - Pan, Ping Yue

PY - 2024/12/20

Y1 - 2024/12/20

N2 - Surface availability of the dopamine (DA) transporter (DAT) critically influences DA transmission. Here, we present a protocol that describes the preparation of mouse ventral midbrain neurons, the expression of a new optical sensor, DAT-pHluorin, and the utilization of this sensor to analyze the surface availability of DAT in live neurons via fluorescent microscopy. This approach allows quantitative measures of basal surface DAT fraction under genetic backgrounds of interest and live trafficking of DAT in response to psychoactive substances. For complete details on the use and execution of this protocol, please refer to Saenz et al.1

AB - Surface availability of the dopamine (DA) transporter (DAT) critically influences DA transmission. Here, we present a protocol that describes the preparation of mouse ventral midbrain neurons, the expression of a new optical sensor, DAT-pHluorin, and the utilization of this sensor to analyze the surface availability of DAT in live neurons via fluorescent microscopy. This approach allows quantitative measures of basal surface DAT fraction under genetic backgrounds of interest and live trafficking of DAT in response to psychoactive substances. For complete details on the use and execution of this protocol, please refer to Saenz et al.1

KW - cell biology

KW - cell culture

KW - cell-based assays

UR - https://www.scopus.com/pages/publications/85207336475

U2 - 10.1016/j.xpro.2024.103358

DO - 10.1016/j.xpro.2024.103358

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JO - STAR Protocols

JF - STAR Protocols

IS - 4

M1 - 103358

ER -

Protocol for live neuron imaging analysis of basal surface fraction and dynamic availability of the dopamine transporter using DAT-pHluorin

Abstract

Keywords

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