Calpain-TRPC6 Signaling Pathway Contributes to Propofol-Induced Developmental Neurotoxicity in Rats

Background Compelling experimental evidence suggests a risk of neuronal damage following early childhood exposure to anesthesia and sedation drugs, including propofol. We investigated whether the transient receptor potential canonical 6 (TRPC6) channel could protect neonatal rats against developmental neurotoxicity following prolonged exposure to propofol. The potential role of calpain, a neuronal TRPC6 protease, was also investigated. Methods Postnatal day 7 rats were exposed to five bolus injections of 25 mg/kg propofol or 10% intralipid at hourly intervals. Acute neuronal injury was assessed by the expression pattern of terminal deoxynucleotidyl transferase nick-end labeling staining and cleaved–caspase-3 in the prefrontal cortex. The Morris water maze test was used to evaluate learning and memory functions in later life. Pretreatments consisting of intracerebroventricular injections of a TRPC6 agonist, TRPC6 inhibitor, or calpain inhibitor were used to confirm the potential role of a calpain–TRPC6 pathway. Results Prolonged exposure to propofol induced acute neuronal injury, downregulation of TRPC6, and enhancement of calpain activity in the prefrontal cortex up to 24 h after anesthesia. It also induced later behavioral disorders, manifesting as longer escape latency and as fewer platform-crossing times and less time spent in the target quadrant during postnatal days 35–42. These propofol-induced effects were attenuated by TRPC6 agonist and calpain inhibitor while exaggerated by TRPC6 inhibitor. Treatment with calpain inhibitor also attenuated the propofol-induced TRPC6 downregulation in the prefrontal cortex. Conclusions A calpain–TRPC6 signaling pathway contributes to propofol-induced acute neuronal injury and long-term behavioral disorders in neonatal rats.

Neuropathology Evaluation in Juvenile Toxicity Studies in Rodents: Comparison of Developmental Neurotoxicity Studies for Chemicals With Juvenile Animal Studies for Pediatric Pharmaceuticals

The developmental neuropathology examination in juvenile toxicity studies depends on the nature of the product candidate, its intended use, and the exposure scenario (eg, dose, duration, and route). Expectations for sampling, processing, and evaluating neural tissues differ for developmental neurotoxicity studies (DNTS) for chemicals and juvenile animal studies (JAS) for pediatric pharmaceuticals. Juvenile toxicity studies typically include macroscopic observations, brain weights, and light microscopic evaluation of routine hematoxylin and eosin (H&E)-stained sections from major neural tissues (brain, spinal cord, and sciatic nerve) as neuropathology endpoints. The DNTS is a focused evaluation of the nervous system, so the study design incorporates perfusion fixation, plastic embedding of at least one nerve, quantitative analysis of selected brain regions, and sometimes special neurohistological stains. In contrast, the JAS examines multiple systems, so neural tissues undergo conventional tissue processing (eg, immersion fixation, paraffin embedding, H&E staining only). An “expanded neurohistopathology” (or “expanded neuropathology”) approach may be performed for JAS if warranted, typically by light microscopic evaluation of more neural tissues (usually additional sections of brain, ganglia, and/or more nerves) or/and special neurohistological stains, to investigate specific questions (eg, a more detailed exploration of a potential neuroactive effect) or to fulfill regulatory requests.