Nomination to NIEHS National Toxicology Program for research on the heritable (germ cell) effects of general anesthetic agents
National Toxicology Program
National Institute of Environmental Health Sciences
US Department of Health and Human Services
November 29, 2018
Re: Nomination for research on the heritable (germ cell) effects of general anesthetic agents
Each year in the United States 1.5 million young children and an unknown number of fetuses are acutely exposed to important and necessary, yet neurotoxic and epigenetically potent chemicals—general anesthetic gases (GA). While there has been ample research on somatic effects of these agents, there has been little attention to germ cell, or heritable, effects of exposure during critical windows of germline development.
This represents a shocking void in the research, given that it is established that GA can cause substantial changes in gene and protein expression, and that animal models have consistently demonstrated intergenerational behavioral and learning impairment. Furthermore, in my experience as a leader in the autism community and as a research philanthropist, I have detected alarming patterns of neurodevelopmental pathology in offspring of parents who were heavily exposed to GA in very early life, during vulnerable periods of gametogenesis.
(1) GA causes epigenetic modification of the genome
It has been known for some time that general anesthetics induce epigenetic modification of the genome. General anesthesia includes agents such as isoflurane, enflurane, halothane, and sevoflurane not only influence neuronal function, they also induce epigenetic alterations such a chromatin changes, histone modifications and shifts in DNA methylation. (Csoka et al., 2009; Vutskits et al., 2018).
GA agents can cause apoptosis and amyloid beta-protein accumulation, and neuronal damage, with potential mechanisms including enhanced protein misfolding and aggregation (Csoka et al., 2009). It has been shown that GA can cause substantial changes in gene and protein expression (Pan et al., 2009; Rampil et al., 2006). For example, even brief exposure to isoflurane leads to widespread changes in genetic control in the amygdala six hours after exposure (Pan et al., 2006). GA can modulate histone acetylation and as such may have deleterious effects on transcription of genes crucial for proper synapse formation and cognitive development (Dalla Massara et al., 2016). A recent study on sevoflurane demonstrated the induction of DNA methylation modification and changes in expression go brain-relevant genes. The investigators found a sex-specific decrease in KCC2 and increased DNA methylation of the KCC2 gene promoter in the sperm of F0 exposed sires (Ju et al., 2018).
Germ cells are at heightened sensitivity to epigenomic error during the early phases of gametogenesis (fetal and neonatal periods) when the DNA is globally demethylated and then remethylated in a sex-dependent manner. Abnormal patterning of DNA methylation or chromatin architecture in a particular region of germline in the developing fetus or neonate could lead to dysregulated somatic development in the generation borne of those exposed germ cells. This is not mere speculation. Empirical evidence supports this idea.
(2) Mammal models demonstrate intergenerational toxicity
[Note F2 = autistic child; F1= parent; F0 = gestating mother of F1 and grandmother of F2.]
The three mammal studies published on the question all point in the same direction. Most recently, neonatal exposure to the widely used general anesthetic agent sevoflurane (sub-clinical doses, in this case) affected the brains and behavior of the next generation of male rats through epigenetic modification of gene expression, while F1 females were at diminished risk (Ju LS, et al. 2018). The implications of this finding could be significant. To quote an accompanying editorial, ““Hence, we are faced with a real possibility that general anaesthetics are not innocuous agents that ‘only put children to sleep’ but rather formidable modulators of chromatin remodeling and function…. The current study extends previous reports of sex differences by showing that anaesthetic exposure itself can alter expression of chloride channels in certain brain regions and that this effect is heritable from exposed male parents to unexposed offspring” (Vutskits et al., 2018).
The two earlier studies to observe offspring impairment are from the 1980s, well before epigenomic or chromatin mechanisms were under consideration. Learning retardation was seen in F2 mouse offspring of F1 parents exposed to general anesthesia in utero—in other words, mental impairment in the grandpups of the exposed gestating dams (Chalon et al., 1981). This phenomenon was also seen with the sires were exposed in adulthood. The general anesthetic agent enflurane administered to male mice was found to adversely affected learning function of their offspring (Tang et al., 1985).
(3) Autism family case studies suggest GA acts as a potent germline toxicant during sensitive periods of gametogenesis
For several years I have been discussing the question of parental fetal or early life exposures with autism parents. After numerous interviews I perceived a strong pattern of F2 pathology where the F1 had an early life exposure to GA, often by virtue of repeated and complicated surgeries.
(a) F0 gestational exposure. These are cases where the F0 grandmother of the F1 parent, male or female, had surgery during gestation with the F1. Reasons for the F0 surgeries during pregnancy included an appendectomy, surgery following an automobile accident, and surgery to correct a problem with the placenta. When an F1 parent had this prenatal exposure, he or she often had multiple F2 offspring, male and female, with autism. From a biological point of view this multiplex phenomenon would make sense because the early germ cells at this stage would likely be similarly exposed.
(b) F1 early childhood exposure. These are cases where the F1 parent, male or female, had surgery or a series of surgeries, in early life. Reasons for the F1 surgeries included tumor removal, hernia repair, surgeries to correct heart defects, and surgeries to correct birth defects such as clefts and club foot. When an F1 parent had this early life exposure, I saw he or she often had multiple F2 offspring, male and female, with autism. From a biological point of view this multiplex phenomenon might also make sense because the female oogonia are undergoing imprinting through the first year and are not yet mature, and the pre-meiotic male spermatogonial stem cells could also retain errors in their later-differentiated spermatocytes.
(c) F1 paternal pre-puberty/puberty exposure. These are cases where the F1 father had a series of surgeries around the time of puberty and beyond. In this category, two stories jump out. I have two male friends who suffered gunshot wounds in late childhood. Both underwent multiple surgeries in puberty and beyond to correct extensive damage. They each have one F2 son with extremely severe autism. Actually the word autism does not do their phenotypes justice, as their conditions are catastrophic, involving profound intellectual disability and severe behaviors, including in one case continuous and extreme self-injurious behaviors. In both of these cases the father also has F2 children who are typically developing. From a biological point of view this simplex phenomenon perhaps make sense because the germ cells were affected at a later stage of differentiation.
I am aware these are all anecdotes and alone prove nothing. But it is worth nothing that in these cases, the families had no history of autism, and to my knowledge, the families and children had no risk factors for autism. As a rough control group, I noticed that where the F1 parent’s siblings did not have these sorts of surgical exposures, the F2s were typically developing.
In spite of the importance of the questions presented, the NTP might respond, “we only study environmental chemicals, and GA falls under the purview of the FDA, not the NIEHS.” In spite of that, I entreat the NTP to include these exposures in the NTP for the following reasons:
The question remains: which agents to study? Because halothane was the most popular GA during the post-war decades and sevoflurane is popular today, I would suggest the NTP begin with those two agents and then expand to others depending on the signals from pilot data. It may turn out that thorough investigation of germline effects (via many different windows of germline development, many different doses, and/or many different mixtures) may well be warranted.
Thank you for your kind consideration of this nomination.
Very truly yours,
Chalon J, Tang CK, Ramanathan S, Eisner M, Katz R, Turndorf H. 1981. Exposure to halothane and enflurane affects learning function of murine progeny. Anesth Analg 60:794–7.
Csoka AB, Szyf M. Epigenetic side-effects of common pharmaceuticals: A potential new field in medicine and pharmacology. 2009. Med. Hypoth. 73;770–780.
Dalla Massara L, Osuru HP, Oklopcic A, et al. General anesthesia causes epigenetic histone modulation of c-fos and brain-derived neurotrophic factor, target genes important for neuronal development in the immature rat hippocampus. 2016. Anesthesiology 124:1311e27.
Heflich R. 2018. Lecture at Environmental Mutagenesis and Genomics 2018 Meeting, “My Forty Years at the U.S. Food and Drug Administration, Robert H. Heflich, U.S. Food and Drug Administration.” Delivered September 24, 2018.
Ju LS, Yang JJ, Morey TE, Gravenstein N, Seubert CN, Resnick JL, Zhang JQ, Martynyuk AE. 2018. Role of epigenetic mechanisms in transmitting the effects of neonatal sevoflurane exposure to the next generation of male, but not female, rats. Brit J Anesth 121:2;406-4168.
Martinez et al., Thyroid hormone influences brain gene expression programs and behaviors in later generations by altering germ line epigenetic information, Mol. Psych. 2018.
Pan JZ, Wei H, Hecker JG, Tobias JW, Eckenhoff RG, Eckenhoff MF. Rat brain DNA transcript profile of halothane and isoflurane exposure. Pharmacogenet Genomics 2006;16:171–82.
Rampil IJ, Moller DH, Bell AH. Isoflurane modulates genomic expression in rat amygdala. Anesth Analg 2006;102:1431–8.
Tang CK, Chalon J, Markham JP, Ramanathan S, Turndorf H. Exposure of sires to enflurane affects learning function of murine progeny. 1985. Obstet. Anesth. Dig. 5:2,67.
Vutskits L, Sall JW, Jevtovic-Todorovic V. A poisoned chalice: the heritage of parental anaesthesia exposure. 2018. Brit. J. Anesth. 121;2,337-339.
Woodcock, J. Letter to Escher in response to citizens petition to withdraw approval for 17-alpha hydroxyprogesterone caproate as a drug used in pregnancy. 2018. http://www.germlineexposures.org/uploads/6/4/0/9/6409433/fda-2015-p-0876.pdf (accessed November 9, 2018).
Jill Escher, Escher Fund for Autism, is a California-based science philanthropist and mother of two children with severe autism, focused on the question of how environmentally induced germline disruptions may be contributing to today's epidemics of neurodevelopmental impairment. You can read about her discovery of her intensive prenatal exposure to synthetic hormone drugs here. Jill is also president of Autism Society San Francisco Bay Area.