**5. Pathogenesis**

The molecular pathogenesis of anesthesia-induced neurotoxicity has also been investigated in experimental studies.

Neonates are born with approximately 100 billion neurons, and the number of neurons does not increase over time. The neonatal brains weigh approximately 300–400 g. Increased myelination, synapse formation, neuron maturation, and proliferation of glial cells increase the weight of the brain to 1100 g at 3 years of age and 1300–1400 g at adulthood. A newborn infant has approximately 50 trillion synapses, increasing to 1000 trillion within the first year of life and decreasing to 500 trillion in adulthood. Critical periods for brain development are the intrauterine period, the first 3 years of life and puberty [38–40].

Thus, brain maturation is not complete at birth, and there is a heterogeneous maturation process in the brain following birth. Maturation is particularly slow in the cortex and in the limbic system [38–40]. Alteration of neurotransmission in the immature brain due to anesthesia exposure may lead to future impairments.

Synaptogenesis has been defined as the most important period of brain development, also described as the "fragile period" or "critical period." Synaptogenesis consists of five phases. The greatest leap in synapse formation occurs in phase 3, which is sometimes referred to as the "big bang." Phase 3 corresponds to the neonatal period. Following phase 3, synaptogenesis continues with the same speed during phase 4. This phase is referred to as the plateau phase, corresponding to infancy and adolescence. During phase 5, which occurs during adulthood, synaptogenesis continues, but it is limited and localized [41]. The initiation, duration, and end of these critical periods (phase 3 and phase 4) are controlled by multiple genetic and epigenetic mechanisms. The brain's sensitivity to environmental stimuli is at maximum during the neonatal and infancy period when synaptogenesis is also maximized [41].

were under the effect of ketamine for 9 or 24 hours. In cell culture study of Bosnjak et al. [25], they demonstrated that ketamine decreases neuronal viability time and dose dependently, leads to neuronal ultrastructural abnormalities, causes depolarization of mitochondrial membrane potential, induces apoptotic pathway, causes cytochrome c release from mitochondria

Yu et al. examined neuroapoptosis and long-term behavioral changes in PND7 rats that were given single and repetitive doses of propofol. Their findings included reduction in neuron density, morphological changes in pyramidal cells, apoptosis, and suppressed release of excitatory neurotransmitters. Additionally, these effects were more pronounced among the

Benzodiazepines (clonazepam, diazepam, and midazolam), which are intravenous anesthetics, have controversial effects on apoptosis; however, barbiturates (pentobarbital, phenobarbital) clearly increase apoptosis. The few studies that have examined the effects of sodium thiopental reported that exposure did not result in increased apoptosis [27–33]. Thompson [34] has suggested high-dose narcotic anesthetic for neonatal and infant. But, fetal and neonatal chronic exposure to opioids has been associated with neuronal changes. Although opioid-based anesthesia and opioids coadministered with inhalation anesthetics have been shown to reduce apoptosis, safety has not been demonstrated with these preparations [35, 36]. However, these studies are controversial and their safety has been in question. Another study has demonstrated that dexmedetomidine, the current intravenous anesthetic, reduces

The molecular pathogenesis of anesthesia-induced neurotoxicity has also been investigated

Neonates are born with approximately 100 billion neurons, and the number of neurons does not increase over time. The neonatal brains weigh approximately 300–400 g. Increased myelination, synapse formation, neuron maturation, and proliferation of glial cells increase the weight of the brain to 1100 g at 3 years of age and 1300–1400 g at adulthood. A newborn infant has approximately 50 trillion synapses, increasing to 1000 trillion within the first year of life and decreasing to 500 trillion in adulthood. Critical periods for brain development are

Thus, brain maturation is not complete at birth, and there is a heterogeneous maturation process in the brain following birth. Maturation is particularly slow in the cortex and in the limbic system [38–40]. Alteration of neurotransmission in the immature brain due to anesthe-

Synaptogenesis has been defined as the most important period of brain development, also described as the "fragile period" or "critical period." Synaptogenesis consists of five phases. The greatest leap in synapse formation occurs in phase 3, which is sometimes referred to as

the intrauterine period, the first 3 years of life and puberty [38–40].

sia exposure may lead to future impairments.

into cytosol, and induces free oxygen radical production.

group that was subject to repeated doses of propofol [26].

prenatal toxicity caused by propofol [37].

**5. Pathogenesis**

106 Current Topics in Anesthesiology

in experimental studies.

Anesthetics elicit their effects by enhancing the activity of major inhibitory neurotransmitters gamma-aminobutyric acid (GABA) and glycine or antagonizing the N-methyl-D-aspartate (NMDA) receptors of the major excitatory neurotransmitter glutamate. During brain development, GABA facilitates cell proliferation, neuroblast migration, and dendritic maturation, and unlike in adults, it acts as an excitatory neurotransmitter during infancy rather than an inhibitory neurotransmitter [42, 43]. This is because these two mediators increase the permeability of the cell membrane to chloride ions through intrinsic chloride-conducting ion pores. After the permeability of the GABA, ligand-gated ion channel to chloride is increased, KCC2 K+/Cl-2 cotransporter aids in influx of chloride ion. Thus, the neuron is hyperpolarized and its activity is suppressed. However, because KCC2 expression is low during the early period of development, the chloride action potential is reversed by GABAA and glycine receptor activity, leading to neuronal depolarization and increased permeability to chloride. Clinical studies have shown that sevoflurane, isoflurane, and propofol cause excitability in electroencephalogram in neonates [44–46]. The major excitatory neurotransmitters glutamate and aspartate are present in the brain at very high concentrations (glutamate 10 mmol/L and aspartate 4 mmol/L). Glutamate and aspartate direct synaptic signaling at nerve terminals and control ion intake to neurons. They have been found to influence synaptogenesis, neuronal plasticity, learning, and memory [47–49]. Although the excitatory neurotransmitters are normally responsible for nerve conduction, they are also potential sources of neurotoxicity. An abnormal decrease in glutamate may disturb normal excitation, and abnormal increases may cause excitotoxicity and cell death by disturbing calcium homeostasis. Glutamate and similar amino acids have been shown to cause acute swelling in the neuron body, dendrites, and glia and also promote neuronal degeneration over extended periods of time. For this reason, there is a delicate mechanism acting in normal conditions to regulate glutamate levels in the synaptic gap involving reuptake of excess glutamate from the synaptic gap through receptors present in presynaptic end of nerve terminal and glial cells. Although glutamate is a strong and rapid-acting toxin under physiological conditions, this mechanism ensures that even direct application to the brain does not cause damage [47]. Nevertheless, pathological conditions that result in insufficiency of this system or cause release of large amounts of glutamate would lead to neuronal loss. For these reasons, anesthesia applications are believed to disrupt the balance between excitatory and inhibitory neurotransmission and thus cause neuronal injury [47–49].

Regarding neuronal viability and development, one of the most studied neurotropins in neonatal subjects is brain-derived neurotrophic factor (BDNF). Mature BDNF is formed by destruction of proBDNF in the synaptic gap by the action of plasmin. Mature BDNF binds to the TrkB receptors present on the postsynaptic membrane and enhances viability of the target cell. However, in conditions where plasmin release is reduced or blocked, such as when anesthesia is applied, proBDNF cannot be converted to the mature form, and it stimulates p75NTR instead of the TrkB receptor. Activation of p75NTR receptor, also called the "death receptor," leads to actin depolymerization and apoptosis. Head et al. [50] demonstrated that isoflurane causes apoptosis in the neonatal mice brain through this pathway.

Apoptosis is a programmed cell death that can occur in both physiological and pathological conditions. Apoptosis is physiologically present in the developing brain, occurring at a rate of approximately 1%. However, apoptosis that occurs following pathological processes like hypoxia and ischemia is typically problematic. Several experimental studies have shown that apoptosis is increased following anesthesia exposure. However, it is not possible to conduct such studies in humans. Therefore, it is difficult to estimate the rate of apoptosis following anesthesia exposure in humans to what extent this apoptosis affects maturation of the developing brain. Experimental studies have shown that anesthesia induces apoptosis via intrinsic and extrinsic pathways. Anesthesia application causes leakage of cytochrome c and translocation of Bax protein to the mitochondria, leading to activation of the apaf-1 and caspase pathways, respectively. This in turn results in lipid peroxidation via release of free oxygen radicals. Apoptosis occurs not only in intrinsic pathway but also in extrinsic pathway which activates Fas protein [51–53].

There are three publications that demonstrate the relationship between microRNA and anesthetic-induced developmental neurotoxicity; according to these publications, while propofol downregulates microRNA-21, ketamine upregulates microRNA-34a, microRNA-34c, and microRNA-124 and downregulates microRNA-137 [54–56].

In cell culture models, it has been demonstrated that neuron development is highly dependent on the actin cytoskeleton, and anesthetics are dangerous for actin regulation [57–59].

Tau protein hyperphosphorylation at serine 404 demonstrates neurodegeneration and is induced by ketamine. Therefore, microtubules are disrupted and damaged [60].

Translocator protein (TSPO, 18 kDa) is a biomarker that could be used for evaluation of reactive gliosis and microglia activity and has the potential for use in noninvasive imaging using positron emission tomography and single photon emission computed tomography [61]. The relationship between anesthesia-associated neurotoxicity and DNA methylation and gene expression has been investigated [62].

Treatment strategies to reduce neurodegeneration induced by anesthetics have also been widely investigated. Lithium, melatonin, estradiol, pilocarpine, dexmedetomidine, xenon, erythropoietin, L-carnitine, hydrogen gas, and pramipexole are among the leading candidates for this emerging therapy [63, 64].
