Introduction

The Ethological Motivated Light/Dark Box was developed and adapted from the traditional Light/Dark Box and traditional Sociability chamber to assess the limitations of scoring and viewing natural ethological rodent behaviors (Neurwirth et al., 2017). The Ethological Motivated Light/Dark Box allows for the study of re-habituation and latency between stranger interactions. This apparatus is targeted at investigating the relationships of the frontal cortex, amygdala, and hippocampal brain structures. This apparatus can be used for the assessment of these brain structures and genetic, epigenetic, or environmental factors in social affective disorders such as ADD and ADHD, Autism, Alzheimer's Disease, Parkinson's Disease, Schizophrenia, and environmentally induced social learning disabilities.

Through understanding the involvement of social motivation and visual recognition during learning, this apparatus addresses how these learning behaviors may interfere with early or late memory formation/encoding and consolidation or reconsolidation in real-time conscious states in social contexts. Moreover, the Ethological Motivated Light/Dark Box can also be used to observe the effect of different pharmacological and genetic manipulations, brain lesions, and more on neurodegenerative and social affective disorders.

The Ethological Motivated Light/Dark Box can be reliably used for visual demonstrations of neuro-social behaviors of rodents and to record real-time accurate data.

Training Protocol

Clean the apparatus after every trial to prevent odor cues from previous trials affecting task performance. Appropriately light the apparatus. A tracking and recording system such as ConductVision or the Noldus Ethovision XT can be used to assist with observations.

Stranger to test rodent combinations can include an old stranger to younger test rodents and vice versa. Gender combinations can also be examined using this apparatus.

Pre-training Protocol.

1. Place the testing room at 6 Lux in illumination. 
2. Stranger introduction: Introduce the “stranger” rodent to the social chamber three times a day for ten minutes.
3. Score nervous behaviors (biting, circling, excessive grooming, etc) of stranger rodents until there is a 70% reduction in these behaviors. The stranger rodent is now ready to use as testable strangers. Use a rotation of 3 strangers during testing.

Testing

1. Testing day 1: Place test rodent in light/ dark box portion of the apparatus, allow for acclimatization for 10 mins.
2. Testing day 2: Place the test rodent in the light/ dark box portion of the apparatus and the stranger rodent in the social chamber. In ten minutes the following parameters can be measured:
   • Time spent at the social hole.
   • Time spent in light / dark chambers.
   • Latency time between social hole interaction and time spent in the dark chamber.
3. For social deficit studies such as in rodent autism models, novel objects can be placed within the social chamber

Literature Review

Investigation of the effect of Taurine administration on learning deficits induced by developmental Pb2+ (lead) exposure in rats 

Neuwirth et al. (2017) investigated the effect of taurine administration on learning deficits induced by developmental lead exposure in Long-Evan rats using the Neuwirth Hole board. Before testing, control rat pups were fed a regular diet while the lead-exposed rats were fed a diet containing 1.5g/kg lead acetate. Twenty-two subjects were randomly assigned to four treatment groups: Control, Control + Taurine, Lead, and Lead +Taurine. Testing on the Neuwirth Holeboard was performed on two consecutive days when the subjects were at postnatal day 30-40. The Taurine groups were administered with 43 mg/kg i.p. injection of Taurine 15 minutes before testing. On the first day, the subjects were allowed to explore the apparatus for 10 minutes to assess their anxiety behaviors. Testing on the second day was performed in the same manner as the first day; however, four Petri dishes, each having a novel olfactory gradient (vanilla, lemon, orange, and almond) were placed below the four corners of the Neuwirth Holeboard apparatus to encourage exploratory behaviors. The results from day 1 indicated that lead exposure increased freezing behaviors. On day 2, lead exposure reduced exploration behaviors as compared to controls. Moreover, Taurine administration showed little effects on control on both testing days; however, taurine administration in the lead-exposed subjects reduced freezing and increased exploration behaviors at similar levels to controls.

Summary

• The Neuwirth Holeboard is an adaptation of the conventional rodent Hole Board and is designed to improve the study of exploratory-versus anxiety-related behaviors in rodents.
• The Neuwirth Holeboard is used to assess anxiety and exploratory behaviors in rodents.
• It is an adaptation of the conventional rodent Hole board. Therefore, it can be used to examine several other unconditioned rodent behaviors as well, such as spatial working memory and spatial learning. 
• It consists of a chamber with a perforated floor containing sixteen 1/2" holes spaced equally apart. Under each hole, contextual cues or food rewards can be placed.
• The Neuwirth Holeboard can test the effect of different drugs or diseases and disorders on exploratory and anxiety-related behaviors of rodents. 

References

1. Neuwirth LS, Volpe NP, Corwin C, Ng S, Madan N, Ferraro AM, Furman Y, El Idrissi A. Taurine Recovery of Learning Deficits Induced by Developmental Pb2+ Exposure. Adv Exp Med Biol. 2017;975 Pt 1:39-55. DOI: 10.1007/978-94-024-1079-2_4. PMID: 28849442.