Early Developmental Exposure to Volatile Anesthetics Causes Behavioral Defects in Caenorhabditis elegans

Abstract

BACKGROUND Mounting evidence from animal studies shows that anesthetic exposure in early life leads to apoptosis in the developing nervous system. This loss of neurons has functional consequences in adulthood. Clinical retrospective reviews have suggested that multiple anesthetic exposures in early childhood are associated with learning disabilities later in life as well. Despite much concern about this phenomenon, little is known about the mechanism by which anesthetics initiate neuronal cell death. Caenorhabditis elegans, a powerful genetic animal model, with precisely characterized neural development and cell death pathways, affords an excellent opportunity to study anesthetic-induced neurotoxicity. We hypothesized that exposing the nematode to volatile anesthetics early in life would induce neuron cell death, producing a behavioral defect that would be manifested in adulthood. METHODS After synchronization and hatching, larval worms were exposed to volatile anesthetics at their 95% effective concentration for 4 hours. On day 4 of life, exposed and control worms were tested for their ability to sense and move to an attractant (i.e., to chemotax). We determined the rate of successful chemotaxis using a standardized chemotaxis index. RESULTS Wild-type nematodes demonstrated striking deficits in chemotaxis indices after exposure to isoflurane (ISO) or sevoflurane (SEVO) in the first larval stage (chemotaxis index: untreated, 85 ± 2; ISO, 52 ± 2; SEVO, 47 ± 2; P < 0.05 for both exposures). The mitochondrial mutant gas-1 had a heightened effect from the anesthetic exposure (chemotaxis index: untreated, 71 ± 2; ISO, 29 ± 12; SEVO, 24 ± 13; P < 0.05 for both exposures). In contrast, animals unable to undergo apoptosis because of a mutation in the pathway that mediates programmed cell death (ced-3) retained their ability to sense and move toward an attractant (chemotaxis index: untreated, 76 ± 10; ISO, 73 ± 9; SEVO, 76 ± 10). Furthermore, we discovered that the window of greatest susceptibility to anesthetic neurotoxicity in nematodes occurs in the first larval stage after hatching (L1). This coincides with a period of neurogenesis in this model. All values are means ± SD. CONCLUSION These data indicate that anesthetics affect neurobehavior in nematodes, extending the range of phyla in which early exposure to volatile anesthetics has been shown to cause functional neurological deficits. This implies that anesthetic-induced neurotoxicity occurs via an ancient underlying mechanism. C elegans is a tractable model organism with which to survey an entire genome for molecules that mediate the toxic effects of volatile anesthetics on the developing nervous system.

The genetics of isoflurane-induced developmental neurotoxicity

“…The plates were divided into 3 regions in a modification of the technique described by Bargmann [3, 18]. One region contained an attractant (a 20ul spot of OP50) while the opposite region contained no attractant.…”

“…These pathological changes were accompanied by a learning defect that persisted into adulthood in the rat. It is now established that, from nematodes [3] to rodents [1, 4, 5] and to primates [6, 7], volatile anesthetics in isolation are capable of inducing neurodegeneration in the developing nervous system [8–11]. It remains unclear how great a risk anesthetic exposure poses to the newborn human at clinical doses and lengths of time [9, 12, 13].…”

“…In this manner, novel and otherwise unrecognized causes of AIN can be discovered and approaches to prevent AIN may be identified. Using changes in chemotaxis, we previously showed that early exposure to the volatile anesthetic isoflurane is neurotoxic to C. elegans [3]. Utilizing a genetic approach, we have identified two pathways, highly conserved from invertebrates to humans, that regulate AIN in the nematode, C. elegans.…”

Isoflurane exposure in newborn rats induces long-term cognitive dysfunction in males but not females

“…However, the safety of anesthesia in this population is a concern and prior retrospective studies have noted a correlation between anesthesia exposure and learning disabilities 1,2 . In addition, various animal models have shown that early anesthetic exposure results in significant long-term behavioral deficits 3-5 .…”

Is a short anesthetic exposure in children safe? Time will tell: a focused commentary of the GAS and PANDA trials

“…Infant rats were given an anesthetic of isoflurane, nitrous oxide and midazolam and were subsequently found to have significant neuronal cell death and lasting memory impairment (10). Since then there have been many animal studies demonstrating this effect in different species (5,11), with different anesthetic agents (12)(13)(14)(15), and affecting different domains of cognition (16)(17)(18). Early studies focused on anesthesia-induced apoptosis (10,19), however the developing brain possesses neurons in excess and normal apoptosis is present during development (20,21).…”