Support for the experiments in awake head-fixed mice

Need help setting up the experiments in awake head-fixed mice? Contact us at mhc-support@neurotar.com for advice on selecting the products and accessories, ensuring their compatibility with other research equipment, using or servicing already purchased devices. But first, take a look at the resources and the frequently asked questions (FAQs) below:

Resources for setting up the experiments in awake head-fixed mice

For selecting the best approach for your research goals, read an overview of approaches for studying the brain functions of non-anesthetized and behaving mice.

An overview of cranial preparations for in vivo imaging and electrophysiology helps to identify the most suitable surgery type. For the surgical implants available from Neurotar, visit the head plates’ page.

The choice of head fixation apparatus follows from the research goals and the choice of cranial preparations. Get oriented with the clamp options available from Neurotar: standard, tilted, Levelt (magnetic), and fUS clamps.

Regardless of the level of your surgical skills, Neurotar’s surgery tutorials (video) for cranial and spinal cord window preparations provide useful tips for optimizing surgical procedures. Due to the tutorials’ sensitive content, we have protected the videos with passwords. No worries, we share the password with all user labs.

Although mousse handling for the Mobile HomeCage follows the standard handling protocols, we recommend reviewing the recommendations for mouse handling in preparation for the experiments in the Mobile HomeCage®.

This page is a must: recommendations for mouse training in preparation for the experiments in the Mobile HomeCage®. We have included tentative timelines for imaging and electrophysiological recordings in awake mice.

Finally, if you are considering testing the Mobile HomeCage® equipment, review the guidelines for the Mobile HomeCage® trials, including the types of trials, their conditions, and a preparation checklist.

FAQ

  • Two-photon microscopy (through cranial window or thinned skull)
  • Electrophysiology in vivo:  patch-clamp, intracellular, extracellular, multi-channel
  • Functional ultrasound (fUS imaging)
  • Optogenetics

High precision tests can be integrated with behavioral monitoring

  • Stable head-fixation of an awake mouse;
  • No general anesthesia during habituation/training or imaging/recording;
  • Efficient stress reduction due to the flat floor and natural cage environment;
  • Natural sensory stimulation (cage floor and walls);
  • Visual stimulation and integration with VR is possible but not required;
  • The mouse controls its own movement;
  • Standard size Mobile HomeCage is compact, compatible with the majority of commercial microscopes;
  • Mobile HomeCage Large is the only head-fixation system that allows using real tangible mazes;
  • The locomotion tracking device (built into the air dispenser) allows monitoring behavior;
  • Cost-efficient.
  • Full-body restraint combined with sucrose or water reward;
  • Air-lifted styrofoam ball (combined with Virtual Reality to compensate for the lack of the sensory input);
  • Linear, circular or cylindrical treadmills (one-dimensional movement only).

These solutions limit the mouse’s control of its movements (granted, this may be desired under certain experimental conditions), and induce significant stress due to unnatural environment.

Alternatives based on the Mobile HomeCage design:

  • AirTrack, primarily a behavioral device. The system is too large for the majority of 2-photon microscopes; stability of head fixation has not been validated for imaging / ephys recordings
  • The Head-fixation apparatus designed by Mark Harnett’s lab (MIT) allows head movement in xy. This marginal improvement must be weight against a significant increase in design complexity and challenges of analyzing the imaging data.

 

Both the standard size Mobile HomeCage and the Mobile HomeCage Large are designed for mice and occasionally used for juvenile rats (<P20) up to 70 g. We are looking for academic partners to continue the work that we have started on adapting the Mobile HomeCage for the research in rats.

The head plates are manufactured from stainless steel and can be reused after soaking overnight in acetone to remove dental cement and cyanoacrylate.

Cranial window implantation surgery (assuming it is done well) does not affect the mouse’s lifespan and health status. We keep the operated animals for up to one year.

We do not recommend implanting a chronic window before P21. Pups must be weaned from their mothers. Acute windows can be done at the neonatal stage. Window implantation in aged animals is feasible; we have operated 10+ months’ old mice for studies on Alzheimer’s disease mouse models.

The most common reason for head plate detachment is poor cleaning of the periosteum from the skull. The grip between the head plate and the skull can be improved in several ways:

  • By carefully removing the periosteum and subsequently drying the skull;
  • By making scratches on the surface of the skull;
  • By priming the skull with a layer of tissue adhesive glue;
  • By using high-quality cement/glue. We use a mixture of Rapid Repair from Panadent and Vetbond or cyanoacrylic glue.

Finally, the loss of a head plate may be caused by insufficient training, and the resulting stress and jerkiness.

Habituation consists of acclimation, handling, and training. For detailed instructions, visit https://www.neurotar.com/mouse-training-for-mobile-homecage/

Mobile HomeCage walls provide a source of natural tactile stimulation. Since mice rely on tactile stimulation more than on their vision, visual stimulation is not necessary in our system. However, it is possible to visually stimulate mice in the Mobile HomeCage. The simplest way to do so is to put stickers on the walls of the Mobile HomeCage. For visual stimulation from a monitor or VR, use a cage with transparent walls or an extra-low wall cage (20 mm wall height). The same extra-low wall cage can be used for stimulating whiskers or delivering odors (if you need to approach the mouse from the front). An alternative option is to mount the stimulation devices on the bridge.

The cage floor and walls may be covered with stickers of different texture, and small objects can be placed on the walls or on the floor. The objects must be light as not to make the cage too heavy and impede its movement.

 

Liquid reward as well as negative reinforcement via air puff are possible in the Mobile HomeCage. We recommend using the lick port and air puff nozzle specially designed to be attached to the Mobile HomeCage clamp: https://www.neurotar.com/product/lick-port-and-air-puff/ The locomotion software allows automating the reward delivery or negative reinforcement for closed-loop experiments in the Mobile HomeCage.

Lick port can also be used to keep the mouse hydrated during long experiments.

The following behavioral paradigms have been validated by the Mobile HomeCage user labs or by Neurotar team:

  • Open field test for assessing baseline levels of motor and exploratory activity and for assessing anxiety;
  • Novel object recognition for assessing memory processing;
  • Spatial memory and learning assessment in T-maze and donut-maze;
  • Social memory and social interactions’ assessment in the sociability maze;
  • The resident-intruder paradigm (modified) upon photostimulation in the hypothalamic aggression locus;
  • Mating behavior (limited to courting behavior, no copulation);
  • Escape response to a looming visual stimulus.

If you don’t find the desired test on the list, do not assume that it cannot be implemented. Be the first to get it done and contact our team to brainstorm about the ways of going about it.

We recommend using a 25x  water immersion objective with high NA (e.g., 1.05). The maximum imaging depth is 700-900 micrometers.

For large diameter objectives, we recommend using the elongated lower lip: https://www.neurotar.com/product/spare-parts/

Please share your objective’s specifications for a more specific recommendation.

Both intra- (whole-cell) and extracellular (cell-attached, field, single unit) recordings are possible in the Mobile HomeCage.  Neurotar team and our customer labs have performed blind patch-clamping in both whole-cell and juxtacellular modes, holding the cells for up to one hour in awake behaving mice. Electrical noise is no greater than with standard patch-clamping in a brain slice setup. Also, 16-channel silicon probes and glass electrodes have been used by several research groups and produced extremely stable, high-quality recordings.

Standard size Mobile HomeCage is compatible with the majority of commercial 2-photon microscopes. Some older microscope models require special parts. We recommend that you share the model and maker of your microscope with our team prior to placing an order.

The Mobile HomeCage Large is compatible with the following commercial 2-photon microscopes without any modifications: Scientifica’s Hyperscope (VivoScope frame), NeurolabwareNikon A1R (gantry frame)Sutter MOMLaVision Biotech’s TriM Scope MatrixFemtonics’ FemtoS-Bridge and Olympus FVMPE-RS (with InnerFocus). With some cage modifications, the large system also works with Bruker’s Ultima Investigator and 2P Plus models, and with Thorlabs’ Bergamo II and DIY 2-photon.

It is possible to use the Mobile HomeCage Large with most commercial microscopes by adding a periscopic extension, such as the Inverterscope available from LSM Tech.

A variety of xy-translation stages have been used with MHC, including Prior’s z-deck and other models capable of carrying the weight of the Mobile HomeCage (6-13 kg, depending on the model) and other accessories, e.g. micromanipulators or reward systems.

We will work with you to ensure that all critical dimensions/clearances are measured prior to purchasing the Mobile HomeCage (e.g., the vertical distance between the objective and xy-stage, the horizontal distance between the objective axis and the microscope frame, etc).

The Mobile HomeCage requires an airflow of 120 L/min; the Mobile HomeCage Large – 150 L/min and the MultiCage Training Arena – 250 L/min. The air pressure requirement is less critical: 0,1-0,2 bar pressure is sufficient.

In most cases, the Mobile HomeCage can operate from a standard air source. If your air source does not satisfy the above requirements, use a portable air pump. Contact us for recommendations. Do not use pressurized air tanks, as they will run out of air very quickly.

Please note that most noise in the system is generated by the air source. The simplest way to reduce noise is to place the Mobile HomeCage away from the air source, e.g., in an adjacent room, or to build an enclosure for the air source.

The Mobile HomeCage is designed to be maximally silent and vibration-free. Acoustic noise produced by the Mobile HomeCage is essentially whisper-level, comparable to a typical laboratory background noise at 35-40dB. When an air pump is used, the overall noise can be somewhat higher but not to the level of being stressful.

Please note that most noise in the system is generated by the air source. The simplest way to reduce noise is to place the Mobile HomeCage away from the air source, e.g., in an adjacent room, or to build an enclosure for the air source.

Use 70% ethanol solution to clean the carbon fiber cage with a soft cloth. Do not use abrasive sponges, acid cleansers, bleach, and other aggressive products. Do not autoclave. The room temperature where the cage is stored must not exceed 30 degrees Celcius.

The foam cages are consumables: while liquids can be wiped off with a paper cloth, neither liquid nor smell can be completely removed. We recommend using individual foam cages and mazes for each animal; it is possible to use the same cage / maze multiple times for the same animal.

OS: Windows 7 64-bit or Windows 10 64-bit. Windows 10 Pro 64-bit is preferred. Incompatible with 32-bit Windows. Not tested for Windows 8.

CPU (desktop): any 4-core, 2015 or newer processor.

CPU (mobile/laptop): any 4-core, 2018 or newer processor.

RAM: at least 4 GB, but 6 GB or more is strongly recommended.

Ports: one type-A USB 3.0 port.

Storage: approx. 120 megabytes per hour of recording.

The latest version of the software can be found on Github: https://github.com/Neurotar/locomotion-tracking-software/releases

We offer free equipment trials, including free shipment back and forth to any location in Europe and North America. For more information visit: https://www.neurotar.com/request-a-mobile-home-cage-trial/ or contact us at mhc-support@neurotar.com