**1. Introduction**

Hypoxic–ischemic (HI) brain injury is a major cause of long-term neurologic disability and death worldwide. Brain damage caused by hypoxia-ischemia responds to a wide variety of factors, being the central nervous system (CNS) especially susceptible to changes in energy levels, mainly glucose concentrations and oxygen [1]. The brain has a 25% glucose and 20% oxygen consumption of total body weight [2, 3]. This high energy demand is attributed to the functions performed by brain cells such as synaptic activity, neurotransmitter recycling and ion transport [2]. Thus, ensuring correct brain metabolism results in optimal neuronal functioning. HI brain injury is mainly characterized by a decrease in the concentration of oxygen and blood flow, which causes an insufficient supply of nutrients to the brain. These pathological conditions lead to cell death due to

the increase in free radical production and depletion of ATP [4]. This phenomenon is observed both in perinatal asphyxia (PA) and in ischemic stroke (IS) [5–7]. Around 15 to 20% of infants that suffer PA will die in the postnatal period and further 25% will develop severe and long-lasting neurological impairments such as cerebral palsy, epilepsy and neurodevelopmental disorders [8], also representing one of the main causes of morbidity in children and adults in the world [9, 10]. Similarly, at a structural level HI injury mainly affects the layers II, III and VI of the cortex, CA1 and CA3 hippocampal areas, striatum and cerebellum [11]. Therefore, the understanding of the underlying mechanisms of this pathology is essential for the establishment of efficient treatments.

Several neuroprotective strategies have been tested, including Selective Estrogen Receptor Modulators (SERMs) and Selective Tissue Estrogenic Activity Regulators (STEARs), which have shown the same benefits as estrogen, including the decrease of reactive oxygen species (ROS), maintenance of cell viability, mitochondrial survival, among others; without its negative side effects [12–14]. However, there are no effective pharmaceutical strategies to prevent the triggering of secondary injury cascades, including oxidative stress and metabolic dysfunction. In this sense, the present chapter summarizes the underlying mechanisms of HI brain injury and compiles several neuroprotective strategies, including SERMs and STEARs.

#### **2. Mechanisms of brain damage in hypoxia-ischemia**

Hypoxia is a condition that affects mainly the brain, and it is characterized by a low concentration of oxygen, affecting the proper functioning of the organs and tissues exposed to it. This insult causes a variety of responses in the brain. An initial response occurs immediately after the insult and is associated with a depletion of ATP, glucose and phosphocreatine inside the brain. This immediate reaction determines the patient's outcome against injury, which in turn triggers a secondary response that occurs several hours later. A temporary energy recovery takes place almost to the initial physiological levels, providing a treatment window between 1 and 6 hours following injury [8, 15, 16]. A third phase of persistent effects lasts for several years [17]. In general terms, global hypoxia affects the cerebral cortex, the sensorimotor cortex, the talamo and the basal ganglia, causing damage in deep gray matter [18].While the complete pathogenic pathways of HI are not fully described, some mechanisms like apoptosis, increased glutamate, calcium overload, mitochondrial dysfunction and oxidative stress have been proposed to contribute to generate neuronal damage [19].

Primary response depends on the energetic failure, which is characterized by the reduction of the energy supply, generating the accumulation of Reactive Oxygen Species (ROS) via lactate production augment, making the cell susceptible to oxidative stress and mitochondrial dysfunction [18]. Besides this, restricted cerebral blood flow causes a switch to anaerobic respiration, reducing ATP and phosphocreatine, and increasing lactic acid production [16]. Low levels of ATP derived from this energetical failure affect the integrity of the cell membrane. Calcium enters easily to the cell causing the membrane depolarization, blocking calcium storage in the cell, which in turn accumulates in the extracellular space. In addition, the ion flux of sodium/potassium is altered by the Na+/K+ pump dysfunction [20]. The second phase of injury is related to the recovery of blood flow and the reestablishment of brain metabolism, characterized by an inflammatory response, excitotoxicity and oxidative stress, being the main responsible for the brain cells death after hypoxia [7, 18].

#### **2.1 Second phase of injury**

Apoptosis as necrosis are the death pathways of the cell. They are present in brain damage caused by hypoxia, being apoptosis the most common death pathway

**211**

*Neuroactive Steroids in Hypoxic–Ischemic Brain Injury: Overview and Future Directions*

in the young brain unchained by mitochondrial failure [21]. Apoptosis can follow two pathways, being the extrinsic triggered by external signals like the tumor necrosis factor alpha (TNF-α), Fatty acid synthase (FAS), and the intrinsic path mediated by internal factors such as DNA damage or cell stress [22]. The extrinsic pathway is involved in the action of caspase 8 and 10, which activate caspase effectors directly, interacting with the intrinsic pathway, and triggering a permeabiliza-

The Intrinsic pathway is mediated by the release of apoptotic factors such as cytochrome-c, Serine protease HTRA2, mitochondrial (Omi/HtrA2), apoptosis inducing factor (AIF), endonuclease G (endoG), Second mitochondria-derived activator of caspase/direct inhibitor of apoptosis-binding protein with low pI (Smac / Diablo) after permeabilization of the membrane. These apoptotic factors can trigger cell death processes that can be mediated by caspase-dependent pathways. Each of these factors has a role in programmed death. Cytochrome c interacts with Apoptosis protease-activating factor-1 (Apaf-1), creating the apoptosome. Smac/Diablo interacts with apoptosis inhibitors, AIF and endoG act through a caspase- dependent pathway. These are translocated to the nucleus, causing nuclear fragmentation [24, 25]. Hence, the permeabilization of the mitochondrial membrane has been proposed as a marker of a point of no return in hypoxic injury.

HI injury triggers responses at both the systemic and cellular levels. When the energy supply is interrupted, excitotoxicity occurs through an uncontrolled release of excitatory neurotransmitters such as glutamate, causing an acute cascade damaging neurons and glial cells at cytoplasmic and mitochondrial levels, and also causing disruption of the BBB [23]. Glutamate activates NMDA receptors, causing the accumulation of Ca ++ and nitric oxide (NO), which in turn cause production of ROS. The increased levels of intracellular calcium in neurons and glial cells in turn results in the activation of calcium-dependent proteases, reactive oxygen species (ROS) production, mitochondrial dysfunction, oxidative stress, cytotoxic edema, lipases and deoxyribonuclease (DNase), and the stimulation of pro-cell death pathways [23, 26, 27].

The balance between the oxidant and the antioxidant levels of the cell is called redox homeostasis. An imbalance in favor of the intracellular level of oxidants results in what is known as oxidative stress. This deregulation occurs mainly in two free radicals, the reactive oxygen species (ROS), and the reactive nitrogen species (RNS) [28, 29]. Oxidative stress plays a major role in the pathophysiology of HI, due to the significant damage to nucleic acids (DNA degeneration), lipids (lipid oxidation), proteins and different organelles such as the mitochondria [7]. There are different sources of free radicals (ROS and RNS) following HI, including mitochondrial electron transport chain (ETC), xanthine oxidase (XO), NADPH oxidases (NOX) and nitric oxide synthase (NOS), and arachidonic acid (12/15 lipoxygenase) [26, 28].

Mitochondria plays a vital role in survival of the different cells of the CNS [30]. It is composed of two membranes, one internal and one external, each with different functions. Within these membranes is the matrix. There are enzymes responsible for the main metabolic processes to produce ATP, such as the Krebs cycle, β-oxidation, as well as the metabolism of aminoacids [31]. Additionally, the mitochondria is involved in moderating processes of death (apoptosis) and biogenesis or

*DOI: http://dx.doi.org/10.5772/intechopen.93956*

tion of the mitochondrial membrane [23].

**2.2 Excitotoxicity**

**2.3 Oxidative stress**

**2.4 Mitochondria**

#### *Neuroactive Steroids in Hypoxic–Ischemic Brain Injury: Overview and Future Directions DOI: http://dx.doi.org/10.5772/intechopen.93956*

in the young brain unchained by mitochondrial failure [21]. Apoptosis can follow two pathways, being the extrinsic triggered by external signals like the tumor necrosis factor alpha (TNF-α), Fatty acid synthase (FAS), and the intrinsic path mediated by internal factors such as DNA damage or cell stress [22]. The extrinsic pathway is involved in the action of caspase 8 and 10, which activate caspase effectors directly, interacting with the intrinsic pathway, and triggering a permeabilization of the mitochondrial membrane [23].

The Intrinsic pathway is mediated by the release of apoptotic factors such as cytochrome-c, Serine protease HTRA2, mitochondrial (Omi/HtrA2), apoptosis inducing factor (AIF), endonuclease G (endoG), Second mitochondria-derived activator of caspase/direct inhibitor of apoptosis-binding protein with low pI (Smac / Diablo) after permeabilization of the membrane. These apoptotic factors can trigger cell death processes that can be mediated by caspase-dependent pathways. Each of these factors has a role in programmed death. Cytochrome c interacts with Apoptosis protease-activating factor-1 (Apaf-1), creating the apoptosome. Smac/Diablo interacts with apoptosis inhibitors, AIF and endoG act through a caspase- dependent pathway. These are translocated to the nucleus, causing nuclear fragmentation [24, 25]. Hence, the permeabilization of the mitochondrial membrane has been proposed as a marker of a point of no return in hypoxic injury.
