Background

Tail vein catheterization in mice is a fundamental technique in pharmacological research, providing precise and reproducible drug delivery for studying drug effects, pharmacokinetics, and pharmacodynamics [1,2]. This method is particularly valuable in neuropharmacological studies, where maintaining stable plasma drug concentrations is crucial for understanding drug mechanisms and therapeutic effects [3,4]. However, traditional tail vein injection methods face significant limitations when applied to long-term studies requiring continuous drug administration, particularly in awake, freely moving animals [5,6], due to technical challenges in catheterization, inadequate fixation methods, and long-duration limitations. Current tail vein catheterization methods primarily rely on single injections [6] or short-term catheter placement under anesthesia, which are insufficient for studies requiring extended drug exposure in conscious animals [7]. Due to technical challenges associated with tail vein catheterization, many researchers have adopted alternative approaches, such as intraperitoneal injection instead of intravenous delivery [8] or single bolus injection instead of continuous stable drug administration [7]. While these alternatives may be technically easier to implement, they introduce significant limitations: intraperitoneal injection exhibits slower and unpredictable drug absorption compared to intravenous delivery [8,9], and a single injection cannot maintain the stable plasma drug concentrations required for pharmacokinetic studies and chronic treatment protocols [8,9]. Repeated injections, while providing some temporal control, introduce significant animal stress that may confound behavioral and physiological measurements [10,11]. Short-term catheterization techniques typically maintain patency for less than 10 min in awake animals, which is insufficient for most experimental protocols requiring extended observation periods [12]. Collectively, these commonly used approaches are suboptimal for experiments requiring sustained, precisely controlled intravenous exposure in awake animals, because they either fail to maintain stable plasma drug concentrations over extended periods (bolus or non-IV routes) or introduce restraint- and handling-related stress (restraint-based infusion or repeated injections). Therefore, there remains a need for a low-stress method that supports continuous tail vein infusion for ≥60 min in awake, freely moving mice.

The technical challenges of long-term tail vein catheterization in awake mice stem from several factors. First, the small size and delicate nature of mouse tail veins make them prone to catheter dislodgement during normal animal movements such as grooming, walking, and standing [12]. Second, traditional fixation methods typically require pharmacological restraint, which introduces stress variables that affect experimental outcomes and animal welfare [13,14]. Third, there is currently no standardized protocol that effectively addresses both the issue of catheter stability during extended periods and animal comfort [11,15].

Developing a reliable long-term tail vein catheterization protocol for awake mice would significantly advance research capabilities across multiple fields [16]. Such a protocol would enable continuous drug delivery during behavioral testing, allow maintenance of stable plasma drug concentrations, and minimize animal stress by maintaining natural movement patterns. This is particularly important for diverse research applications, including neuropharmacological research, metabolic studies, behavioral assessments, and neuroimaging studies, where the effects of drugs on behavior, cognition, and neural function need to be evaluated in awake animals without the confounding effects of anesthesia or restraint stress.