Combining Cardiovascular, Respiratory and Neurobehavioral Endpoints for Efficient Study Design
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The document discusses the integration of cardiovascular, respiratory, and neurobehavioral endpoints in safety pharmacology and inhalation toxicology studies, emphasizing the use of Allay restraint technology and models for efficient study design. It presents outcomes from pilot studies on the effects of substances like caffeine and chlorpromazine on these physiological systems in rats, highlighting novel methodologies for monitoring multiple endpoints simultaneously. The findings suggest varying impacts on heart rate, respiratory rate, and body temperature, with an emphasis on the necessity of acclimation and evaluation of device effectiveness in trial setups.
![• 4x4 Latin square cross-over design [7 days between each treatment]
• Male rats (CRL:WI(HAN)) instrumented with telemetry devices for the measurement of
arterial pressure, heart rate, ECG, respiratory rate, respiratory pressure, activity, and body
temperature
• Each rat received oral doses of vehicle (distilled water) and caffeine at 3, 12, and 24 mg/kg
• Each rat received oral doses of vehicle (distilled water) and chlorpromazine at 2, 8, and 16
mg/kg
• Arterial pressures, ECG waveforms, respiratory rate, respiratory pressure, and activity were
monitored continuously for 2 hours prior to dosing and up to 24 hours post-dose
Study Design](https://image.slidesharecdn.com/combiningcardiovascularrespiratoryandneurobehavioralendpointsforefficientstudydesigndsi-161020173055/85/Combining-Cardiovascular-Respiratory-and-Neurobehavioral-Endpoints-for-Efficient-Study-Design-41-320.jpg)
Combining Cardiovascular, Respiratory and Neurobehavioral Endpoints for Efficient Study Design
- 1. Combining Cardiovascular, Respiratory and Neurobehavioral Endpoints for Efficient Study Design Brian M. Roche PhD, DSP, DABT Director of Safety Pharmacology, Charles River Laboratories Jason D. Payseur Senior Scientist – IVIVT, MSD Safety Pharmacology US GlaxoSmithKline
- 2. InsideScientific is an online educational environment designed for life science researchers. Our goal is to aid in the sharing and distribution of scientific information regarding innovative technologies, protocols, research tools and laboratory services. JOIN FOR FREE AT WWW.INSIDESCIENTIFIC.COM
- 3. Combining plethysmography, telemetry and Allay™ restraint technology for the use in safety pharmacology and inhalation toxicology environments Brian M. Roche PhD, DSP, DABT Director of Safety Pharmacology, Charles River Laboratories, Ashland, OH Copyright 2016 B.M. Roche, Data Sciences International and InsideScientific. All Rights Reserved.
- 4. Agenda 1. Allay Restraint Technology • Overview of device • Acclimation process 2. Allay restrainers vs. nose only cone restraint tubes • Acclimation data • Hemodynamic telemetry data 3. Plethysmography and Telemetry: Allay vs Head Out • Hemodynamic assessment • Respiratory function • Resistance and compliance 4. Dosimetry approaches with Allay plethysmography 5. What we’ve learned from conducting this pilot study
- 5. Allay Restraint Acclimation The Process
- 6. Allay Restraint Technology 1 Allay restraint tube 2 Neck clip 3 Plethysmography bell 4 Nose seal1 2 3 4
- 7. Important to allow for adequate time prior to study to acclimate the rats to the restraint device Introducing the allay restraint tube and the rat either through placing the restraint tube or “sled” portion of the device into the home cage and/or manually placing and holding the rat inside the device. Allay Restraint and Acclimation
- 8. After the initial familiarity with the device, the next step was to introduce the restraint portion of the tube. This was conducted by manually placing and holding the rat in the device without the neck clip. Allowing the rat to extend its head and neck out through the front of the device while it explored its surroundings. Allay Restraint and Acclimation
- 9. The next step is to determine the appropriately sized “clip” for the size of the rat’s neck. This process starts as an extension of the manual restraint of the rat within the device. Each time the rat extends it’s neck through the “clip” area, slowly place the neck clip over the rat’s neck. This step requires patience as restraining the rat too early may set you back a step in acclimation. Allay Restraint and Acclimation
- 10. Once the appropriate sized neck clip is in place the rat will begin acclimating to this step of the procedure. Allay Restraint and Acclimation
- 11. Once the rat has been introduced and the appropriate neck clip has been determined, acclimation to the allay restraint technology for respiratory function beings. Depending upon your study design, the next step in acclimation is to add the plethysmography bell. This step is required if collecting respiratory function. At this point the restraint is similar to a head out chamber without a neck seal. This step is critical in the acclimation process for tail placement and temperature regulation. Allay Restraint and Acclimation
- 12. Nose and mouth seal. The nose seal can also be placed on the restraint tube without the plethysmography bell for inhalation delivery of test articles/chemicals. The rat will react once the face whiskers are stimulated but will calm down. Additionally, the nose will be positioned directly in the center of the nose seal. Allay Restraint and Acclimation
- 13. Putting it all together. The device is functioning as a nose only plethysmography chamber for inhalation delivery. This piece is also critical for placement on the inhalation tower. The acclimation will be conducted in steps, with increasing duration. Once the entire Allay restraint device is assembled around the rat, start building up acclimation time to cover the time that the rat will be restrained on study. Allay Restraint and Acclimation
- 14. Acclimation: Allay restrainers vs. nose only restraint cone HD-S21 Telemetry – Hemodynamic data
- 15. Allay Restraint vs. Nose Only Restraint Cone
- 16. Heart rate reduced initially in Allay restrainers
- 17. No change in blood pressure between the two methods of restraint
- 18. EVERY STEP OF THE WAY16 Slight increase in body temperature, expected with no bias flow and tail enclosed in Allay restraint
- 19. Plethysmography and Telemetry: Allay vs. Head Out Restraint Hemodynamic Assessment, Respiratory Function, Resistance and Compliance
- 20. Allay Restraint vs. Head Out Plethysmograph
- 21. Reduction in heart rate through first 60 minutes for Allay restraint Allay Head Out
- 22. No change in systolic blood pressure Allay Head Out
- 23. Slight reduction in diastolic blood pressure for initial 60 minutes in Allay restraint Allay Head Out
- 24. Slight reduction in mean arterial blood pressure for Allay restraint Allay Head Out
- 25. Slight reduction in body temperature for the Allay restrain Allay Head Out
- 26. Decrease in respiratory rate for Allay restraint Allay Head Out
- 27. Increase in tidal volume with Allay restraint Allay Head Out
- 28. No change in minute volume Allay Head Out
- 29. No change in lung compliance Allay Head Out
- 30. No change in lung resistance Allay Head Out
- 31. Dosimetry Approaches With Allay Plethysmography Inhalation drug delivery
- 32. Standard fit for the Allay restraint system with a conventional nose only inhalation system. Inhalation tower designed with a support ring to handle the size and weight of the Allay restraint. Allay Technology and Inhalation Tower
- 33. Telemetry hemodynamic and intrathoracic pressure data were collected in combination with respiratory function by placing a DSI RPC-1 small animal receiver in close proximity to the animal instrumented with HD-S21 transmitter. Adequate separation of the animals throughout the inhalation tower, reduces the risk of cross-talk. Allay Restraint and Telemetry
- 34. What we’ve learned from conducting this pilot study…
- 35. Pros… Ability to combine inhalation and dosimetry Ability to dose via inhalation (nose only cone) Ability to collect dosimetry (head out chamber) Potentially less stressful (nose cone and head out) Plethysmography - no thoracic compaction of rat vs head out Tight seal of device when mounted on inhalation tower Spacing collar for proper placement of head chamber and nose seal No physical obstruction against the throat (head out chamber) Dual chamber (reference) reduces variability of ambient pressure changes in the collection environment (head out chamber)
- 36. Cons… Nose seal o Destruction by rat o Ability to replace nose seal (loss of data) Acclimation process is time consuming Design o Heavy (ring design of inhalation tower) o Rolls (acclimation) o Position of pneumotach and transducer ports (Obstructed by tail/urine/feces)
- 37. Acute Respiratory Disorders • Respiratory Depression • Respiratory Syncytial Virus (RSV) • Acute Respiratory Distress Syndrome (ARDS) • Mucociliary Clearance and Dysfunction • Pneumonia • Cough • Tuberculosis (TB) • Bronchiolitis/ Bronchitis COPD • Emphysema • Chronic Bronchitis • Model Development • Treatment Assessment Asthma • Model Development • Treatment Assessment Lung Fibrosis • Pulmonary Fibrosis • Cystic Fibrosis Safety Assessment • Safety Pharmacology • Toxicology Select any of the links below to learn more about Respiratory Solutions offered by DSI. www.datasci.com
- 38. Combined Cardiovascular, Respiratory, and Neurobehavioral Telemetry Model in the Conscious Rat: Jason D. Payseur Senior Scientist – IVIVT, MSD Safety Pharmacology US GlaxoSmithKline Copyright 2016 J.D. Payseur, Data Sciences International and InsideScientific. All Rights Reserved. A Novel Approach to Study the Acute Physiological Effects of Caffeine and Chlorpromazine Following Oral Administration
- 39. Background • Rodent studies historically focused on a single physiological system • Integration of cardiovascular, respiratory, and neurobehavioral assessments would allow for more insight into how changes in one physiological system impact the others • Advances in recent years have allowed combined models in large animals (i.e. JET/EMKA jacketed models) • The release of a dual pressure catheter in a small animal model implant (model HD-S21) by Data Sciences International (DSI) opens up an opportunity to test a combined rodent model
- 40. Surgical Procedure • Prior to study, rats were implanted with a telemetry device (Model No. HD-S21, Data Sciences International (DSI), St. Paul MN). • 1 pressure catheter was inserted into the abdominal aorta and advanced to a position caudal to the renal bifurcation. Another pressure catheter was inserted under the serosal surface of the esophagus in the thoracic cavity.
- 41. • 4x4 Latin square cross-over design [7 days between each treatment] • Male rats (CRL:WI(HAN)) instrumented with telemetry devices for the measurement of arterial pressure, heart rate, ECG, respiratory rate, respiratory pressure, activity, and body temperature • Each rat received oral doses of vehicle (distilled water) and caffeine at 3, 12, and 24 mg/kg • Each rat received oral doses of vehicle (distilled water) and chlorpromazine at 2, 8, and 16 mg/kg • Arterial pressures, ECG waveforms, respiratory rate, respiratory pressure, and activity were monitored continuously for 2 hours prior to dosing and up to 24 hours post-dose Study Design
- 42. Dose dependant increase in heart rate at all dose levels up to 2 hours post dose. AbsoluteDataChangeFromPre-dose Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 HeartRate (beats/minute) 250 300 350 400 450 500 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 HeartRate (beats/minute) -50 0 50 100 150 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) 24mpk: 67.8 bpm; 19.3% 12mpk: 18.7% 3mpk: 10.2% Effect of Caffeine on Heart Rate
- 43. Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 RespiratoryRate (breaths/minute) 30 60 90 120 150 180 210 240 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 RespiratoryRate (breaths/minute) -50 -25 0 25 50 75 100 125 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) 24mpk: 61.9 bpm; 54.9% 12mpk: 30.0% 3mpk: 22.3% Dose dependant increase in respiratory rate at all dose levels up to 2 hours post dose. AbsoluteDataChangeFromPre-dose Effect of Caffeine on Respiratory Rate
- 44. Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 RespiratoryPressure (mmHg) -10 -5 0 5 10 15 20 25 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 RespiratoryPressure (mmHg) -6 -4 -2 0 2 4 6 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) No statistically significant changes in pressure at any dose level. Effect of Caffeine on Respiratory Pressure AbsoluteDataChangeFromPre-dose
- 45. Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 Activity (AUC) 0 2000 4000 6000 8000 10000 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 Activity (AUC) -4000 -2000 0 2000 4000 6000 8000 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) 24mpk: 400% 12mpk: 489% 3mpk: 257% Significant increases in activity in a dose dependant manner up to 3 hours post dose. Effect of Caffeine on Activity AbsoluteDataChangeFromPre-dose
- 46. Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 Activity (AUC) 0 2000 4000 6000 8000 10000 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 Activity (AUC) -4000 -2000 0 2000 4000 6000 8000 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) 24mpk: 400% 12mpk: 489% 3mpk: 257% Significant increases in activity in a dose dependant manner up to 3 hours post dose. Effect of Caffeine on Activity AbsoluteDataChangeFromPre-dose
- 47. Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 BodyTemperature (DegC) 32 34 36 38 40 42 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 BodyTemperature (DegC) -5 -3 -1 1 3 5 Vehicle Caffeine (3 mg/kg) Caffeine (12 mg/kg) Caffeine (24 mg/kg) Small increases in body temperature in the mid and high doses up to 2 hours post dose, most likely due to the large increase in activity at those times. Effect of Caffeine on Body Temperature AbsoluteDataChangeFromPre-dose
- 48. Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 HeartRate (beats/minute) 250 300 350 400 450 500 Vehicle Chlorpromazine (2 mg/kg) Chlorpromazine (8 mg/kg) Chlorpromazine (16 mg/kg) Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 HeartRate (beats/minute) -50 0 50 100 150 Vehicle Chlorpromazine (2 mg/kg) Chlorpromazine (8 mg/kg) Chlorpromazine (16 mg/kg) 16mpk: 66.1 bpm; 54.9% 8mpk: 66.7% An unexpected increase in heart rate in both the mid and high doses for up to 8 hours post dose (the duration of the light cycle after dosing). Heart rate was comparable to vehicle once lights went out and rats became more active. Effect of Chlorpromazine on Heart Rate AbsoluteDataChangeFromPre-dose
- 49. Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 RespiratoryRate (breaths/minute) 30 60 90 120 150 180 210 240 Vehicle Chlorpromazine (2 mg/kg) Chlorpromazine (8 mg/kg) Chlorpromazine (12 mg/kg) Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 RespiratoryRate (breaths/minute) -50 -25 0 25 50 75 100 125 Vehicle Chlorpromazine (2 mg/kg) Chlorpromazine (8 mg/kg) Chlorpromazine (12 mg/kg) No changes in respiratory rate at any dose level. Very unexpected! Effect of Chlorpromazine on Respiratory Rate AbsoluteDataChangeFromPre-dose
- 50. Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 RespiratoryPressure (mmHg) -10 -5 0 5 10 15 20 25 Vehicle Chlorpromazine (2 mg/kg) Chlorpromazine (8 mg/kg) Chlorpromazine (12 mg/kg) Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 RespiratoryPressure (mmHg) -6 -4 -2 0 2 4 6 Vehicle Chlorpromazine (2 mg/kg) Chlorpromazine (8 mg/kg) Chlorpromazine (12 mg/kg) No significant changes in respiratory pressure immediately post dose, though we do start to see some depressed pressures during the night cycle (hours 9-21 post dose), especially in the high dose. Effect of Chlorpromazine on Respiratory Pressure AbsoluteDataChangeFromPre-dose
- 51. Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 Activity (AUC) 0 2000 4000 6000 8000 10000 Vehicle Chlorpromazine (2 mg/kg) Chlorpromazine (8 mg/kg) Chlorpromazine (12 mg/kg) Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 Activity (AUC) -4000 -2000 0 2000 4000 6000 8000 Vehicle Chlorpromazine (2 mg/kg) Chlorpromazine (8 mg/kg) Chlorpromazine (12 mg/kg) 16mpk: -231% Once again we see no changes in activity until the night cycle, were we see some depression in the high dose. Effect of Chlorpromazine on Activity AbsoluteDataChangeFromPre-dose
- 52. Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 BodyTemperature (DegC) 32 34 36 38 40 42 Vehicle Chlorpromazine (2 mg/kg) Chlorpromazine (8 mg/kg) Chlorpromazine (16 mg/kg) Time After Dosing (Hours) 0 2 4 6 8 10 12 14 16 18 20 22 24 BodyTemperature (DegC) -5 -3 -1 1 3 5 Vehicle Chlorpromazine (2 mg/kg) Chlorpromazine (8 mg/kg) Chlorpromazine (16 mg/kg) A dose dependant decrease in core body temperature from hours 2-14 post dose. Effect of Chlorpromazine on Temperature AbsoluteDataChangeFromPre-dose
- 53. Results (Caffeine) • Rats dosed with 24 mg/kg of caffeine showed the expected transient increase in heart rate (67.8 bpm; 19.3%) as well as increases in respiratory rate (61.9 bpm; 54.9%), and activity (400%). There was no apparent change in respiratory pressure. • Rats given 3 and 12 mg/kg of caffeine also showed increases in heart rate (10.2%; 18.7%), respiratory rate (22.3%; 30.0%), and activity (257%; 489%) in a dose responsive manner • Rats given chlorpromazine showed an increase in heart rate (66.1 bpm; 19.9%) and the expected decrease in activity (- 231%) at a dose of 16 mg/kg. • There was no apparent change in respiratory parameters following administration of chlorpromazine at any dose. Results (Chlorpromazine)
- 54. Conclusions… • Surgically feasible model • Continuous 24 hour respiratory monitoring (time consuming) • Reliable cardiovascular and neurobehavioral results for both caffeine and chlorpromazine • Expected changes in respiratory parameters seen in caffeine dosed rats • Unexpectedly, a decrease in respiratory parameters was not seen in chlorpromazine treated rats, thus further studies must be conducted to determine if the this combined model is able to detect decreases in respiratory rate with confidence.
- 55. Acknowledgements & References • Dennis J. Murphy • Safety Pharmacology staff at GlaxoSmithKline • All studies were conducted in accordance with the GSK Policy on the Care, Welfare and Treatment of Laboratory Animals and were reviewed by the Institutional Animal Care and User Committee either at GSK or by the ethical review process at the institution where the work was performed. 1. Sgoifo, A., et al., “Electrode positioning for reliable telemetry ECG recordings during social stress in unrestrained rats.” Physiology and Behavior. 1996 Mar; 60(6):1397-1401. LINK 2. Lynch III, J. et al., “Comparison of methods for the assessment of locomotor activity in rodent safety pharmacology studies”, Journal of Pharmacological and Toxicological Methods, 64 (2011) 74-80. LINK
- 56. Thank You: Brian M. Roche PhD, DSP, DABT Director of Safety Pharmacology, Charles River Laboratories Jason D. Payseur Senior Scientist – IVIVT, MSD Safety Pharmacology US GlaxoSmithKline For additional information on the solutions presented in this webinar please visit www.datasci.com