Background

Stress is common and inevitable. Severe or chronic stress can result in an array of cognitive, emotional, and physical problems. To understand the molecular, cellular, and physiological underpinnings of stress and its influence on brain function, stress must be modeled in the laboratory. It is conventionally accepted that chronic stress leads to adverse health outcomes while acute stress can be benign or advantageous. For example, chronic stress induced by unpredictable intermittent restraint impaired spatial memory (Peay et al., 2020), whereas one hour of acute restraint stress enhanced novel object recognition memory in male rats (Brivio et al., 2020). However, previous approaches to acute stress typically imposed a single or “simple” stress. Notably, acute stressful life events such as mass shootings or natural disasters may only last minutes to hours, yet consist of physical, emotional, and social stresses occurring at the same time. These events are now being shown to provoke negative long-term outcomes such as PTSD in a significant number of individuals (North et al., 1994; Lowe and Galea, 2017; Musazzi et al., 2017; Novotney, 2018). Therefore, there is an unmet need to model MAS in the laboratory to discover the mechanisms by which they promote negative outcomes.

We have developed a multiple concurrent acute stresses (MAS) paradigm (Figure 1) to examine cognitive and “emotional” impairments resulting from these types of stress (Maras et al., 2014). MAS utilizes mild to moderate stressors: restraint, awareness of peer discomfort, bright lighting, loud noise, and jostling, but delivers these to the animal simultaneously within as little as one hour. MAS lasting two-hours has lasting impact on memory and hippocampal spine integrity, assessed via spatial memory tests, slice physiology, and several quantitative methods to determine spine density and loss (Maras et al., 2014; Chen et al., 2016). For example, adult male mice subjected to restraint only stress or noise only stress, paradigms that are more commonly used in labs to model acute stress, did not have these profound memory deficits. MAS activates additional and distinct brain networks compared to single acute stressors, promoting cross-correlated activation of amygdala and the bed nucleus of the stria terminalis (Maras et al., 2014). These results highlight that MAS may employ different mechanisms to impact brain function compared to commonly utilized acute stressors. Thus, MAS may provide a more optimal model for concurrent acute life stresses that produce profound, negative outcomes.

In contrast to simple acute stress, chronic stress may promote neuropsychiatric pathology. Several protocols have been developed to model chronic stress in rodents. Repeated physical restraint in a conical tube or similar restraining device is a popular paradigm (Kim and Han, 2006). To avoid potential habituation to a single stressor over an extensive period of time, the unpredictable chronic mild stress procedure [UCMS or CUMS, with variations including chronic unpredictable stress (CUS or UCS), chronic mild stress (CMS), and chronic variable stress (CVS)] subjects rodents to several weeks of various stressors, such as noise, restraint, bedding deprivation, and light cycle disruption (Willner, 1997 and 2017; Burstein and Doron, 2018). While this protocol is widely used, it is quite labor intensive and some strains of mice, including the commonly used C57BL/6 strain, can be resistant to the effects of UCMS unless the duration of the protocol is greatly extended (Monteiro et al., 2015).

To circumvent issues with some chronic stress protocols and also engage the same networks as those activated in a single exposure to MAS, the MAS paradigm has been adapted to chronic applications. Repeated multiple concurrent stressors (RMS) for one hour a day across 10 days in adolescent, male C57BL/6 mice induce the loss of sensory and retrosplenial cortical input to the posterior parietal cortex as well as disrupt working memory performance (Libovner et al., 2020). This extensive circuit disruption is not observed in mice subjected to the same amount of restraint only stress, highlighting the specific mechanisms engaged during exposure to MAS.

The MAS protocol facilitates the investigation of the consequences of stress that may be limited in duration, such as natural disasters or mass shootings. This paradigm can further be adapted to chronic applications, capitalizing on the network activation of a single exposure that may not be similarly engaged in other chronic stress paradigms. Furthermore, this protocol is technically simple and titratable, promoting robust and reproducible results.


Figure 1. Multiple Concurrent Acute Stresses Paradigm. Mice are subjected to physical, social, and emotional stresses concurrently through restraint, crowding, jostling, loud noise (e.g., music or random, high frequency beeps), and bright lights.