For aquatic species, temperature is a critical environmental component that can have a big influence on fish aggressive behavior (44). The study results revealed that African catfish exposed to heat stress showed significant increase in aggressive behavior compared to all other groups. This result agreed with Kua et al (45) and Lopezet et al (46). High temperature has been reported to accelerate the metabolic rate of ectothermic species and encourage increased aggression and activity (47). In addition, it is commonly known that cortisol levels and/or neurotransmitter activity influence fish behavior (48). As a result, due to modifications in the stress physiology machinery, alterations in fish behavior might be observed at higher temperatures, which might influence populations in a cascade manner. Neurotransmission alterations have the potential to modify fish social behavior, impacting both intra- and interspecies interactions and potentially influencing ecosystem functioning in warmer climates (49, 50). At the same time, greater temperatures have often been associated with increased locomotor activity, boldness, and aggressive behavior in fish (51, 52). This suggested that these changes allow fish to get higher access to sources of food (to compensate increased metabolic rates), however, this also increases their susceptibility to predators (52).
Analyzing hematological indicators is crucial for assessing fish health in a variety of stressful conditions (53). Because blood components are so sensitive to temperature, any physiological fluctuation will be expressed in the criteria for different blood characteristics (54). In the present study, significant decrease in Hb, RBC and PCV in African catfish exposed to heat stress only compared to other groups. These results agree with the findings of previous studies (55, 56). This could have happened because of the hematopoietic system failure in a stressful environment induced by a high temperature (55). These levels may have decreased due to shrinkage of RBC from thermal stress or comparatively increased erythrophagocytosis of damaged RBC (57). In ZN group there are significant decreases in RBCs, Hb, and PCV compared to control group and this result was similar to Faiz et al (24). The depletion in RBCs of African catfish fed ZnO supplemented diet might result from hemolysis caused by RBCs swelling. Hemolysis of erythrocytes has additionally been documented in Heteroclarias sp (58, 59) and rainbow trout (60). Similarly, according to Kori-Siakpere et al (58), The decrease in erythrocyte indices is associated with anemic condition.
WBC counts fluctuate in all vertebrates, including fish, in response to different stresses such as diseases and chemical pollutants (61). The results revealed a significant decrease in WBCs count in HS and ZN groups compared to CON and ZNH groups. In case of ZN group, other scientists also noted that the number of WBCs had decreased. in Clarias and “Heteroclarias” species in response to Zn (58, 59). The decreased in number of white blood cells (leukopenia) in the present study or previous studies might either come from the bioaccumulation of zinc in various organs, which is toxic and affects cell production from spleen (62) or because of an elevated amount of corticosteroid hormones, which are crucial for the healing and prevention of inflammation (63).
The decrease in WBCs in HS group also reported in different studies (64, 65). In fish, the hypothalamic-pituitary-interrenal and hypothalamic-chromaffin axis are activated in response to environmental stress. Activation of this pathway results in elevated levels of catecholamines, cortisol, glucose, and adrenocorticotropic hormone (66, 67). The non-specific cellular response of tilapia O. mossambicus diminished when it was moved from 27°C to 19 and 35°C. This implies that cortisol and catecholamine, which act as neuro-regulators, may increase, and inhibit immunity (65).
There is a significant increase in neutrophils and monocytes percentage in HS groups while in the same group there is significant decrease in lymphocyte percentage compared to other groups. The same results were reported by Abdel-Ghany et al (68). Moreover, thermal stress results in hypoxia or anoxia (69). In turn, it was found that in red tilapia, hypoxia causes a decrease in lymphocyte counts and an increase in neutrophil and monocyte counts (68, 70). This could be associated with the high level of cortisol (70). Additionally, stress hormones prevent lymphocytes from proliferating (71), granulocytes from undergoes apoptosis (72), and neutrophils and monocytes from emigration from the hematopoietic tissue of the head of kidney into the peripheral blood (73).
The neuroendocrine system of fish can be impacted by elevated stress hormone levels, and some environmental stresses can directly impact fish neurotransmitters (74). Acetylcholine (Ach) is one of several neurotransmitters that is linked to cognitive functions through activation of muscarinic and cholinergic receptors; Acetylcholinesterase (AChE) catalyzes ACh breakdown to keep ACh levels adequate (75, 76). AChE is a commonly used enzyme in fish that is considered a dependable biomarker for assessing environmental stresses, and when exposed to stress, its activity is often inhibited (77). Heat negatively affects the affinity of AChE for acetylcholine, a physiological characteristic that causes sensitivity in fish. Moreover, fish at higher water temperatures have larger metabolic needs, which are not satisfied by an inadequate oxygen supply (78). In this study, the AChE activity in blood of African catfish was significantly inhibited upon exposure to high water temperature (HS group). This was agreed with other studies, (79) which reported that the damselfish (Acanthochromis polyacanthus) significantly reduced its cholinesterase (ChE) activity in response to high temperature stress. Similarly, Kumar et al (80) reported that Acetylcholine activity in the snakehead murrel (Channa striatus) was decreased by high water temperatures. In ZNH group there was a significant improvement in AChE level compared to HS group by using nano-zinc oxide in the diet of African catfish (30mg/kg diet). Similarly, Kumar et al (81) revealed that there was an enhanced level of AChE in fish fed dietary Zn-NPs at 10 mg kg− 1 and exposed to high temperature and lead poisoning. This might have occurred because of synaptic vesicles carrying neurotransmitters being created and exocytosed (81, 82). Also, may be because zinc is a crucial part of the brain and central nervous system, it also has a role as a neuro-secretory product or co-factor in glutamatergic neurons in the fish forebrain (83).
As is widely known, environmental stressors result in the elevation of cortisol (84, 85). Similar findings were obtained in the current study. The blood cortisol was elevated in the group exposed to high temperature (HS group). The hypothalamus-pituitary-interrenal (HPI) and hypothalamic-sympathetic-chromaffin (HSC) pathways are activated during the primary stress response, releasing catecholamines (dopamine, adrenaline, and nor-adrenaline) and corticosteroids into the bloodstream. Corticotropin releasing hormone (CRH) stimulates the pituitary, releasing adrenocorticotropic hormone (ACTH) and melanophore stimulating hormone (MSH) into the bloodstream, which is secreted by the hypothalamus. Additionally, fish's head kidney's chromaffin and interrenal cells releases catecholamines and cortisol (86).
Furthermore, in ZNH group an alleviative role of nano-zinc oxide present by significantly drop the level of cortisol compared with HS group. Linet et al (87) showed that significantly decrease cortisol levels in common carp exposed to abiotic stress by using Zn-NPs. Similarly, When Pangasianodon hypophthalmus were raised in high temperatures and lead (Pb) toxicity, the cortisol level was significantly decreased by supplementing with zinc nanoparticles (10 mg kg− 1 ) (88). Which may be caused by Zn-NPs penetrating the blood-brain barrier and having a positive impact on fish adrenal glands (88, 89). Furthermore, zinc's anti-oxidative effects on cortisol release may possibly be the reason for the results that were observed (90).
The liver's release of metabolites and enzymes is also disturbed when liver tissue is disturbed (91). Any rise in the activity of liver enzymes, such as AST and ALT, is a biological indicator of damage to the liver (92, 93). In this study, blood ALT and AST activities were higher in HS and ZNH groups compared to CON and ZN groups this suggesting that the liver of African catfish had suffered damage to some extent due to heat stress. This was agreed with Dalvi et al (94) which showed elevated AST and ALT activities in catfish (Horabagrus brachysoma) exposed to high temperatures for 30 days. The increased AST and ALT activities at higher temperatures suggest the mobilization of free amino acids for energy production. Similar observations have been reported in C. carpio (95) in response to thermal acclimation. Also, the elevation of liver enzyme activities may indicate enzyme leakage across damaged plasma membranes and/or increased synthesis of liver enzymes by the action of stress (96).
Alkaline phosphatase (ALP) is a crucial metabolic regulator of enzymes in vivo that is directly involved in the metabolism of calcium and phosphate as well as the transfer of phosphate groups. It plays a significant role in the process of nutrient utilization and absorption in aquatic species. ALP can also raise the body's resistance to disease by altering the pathogen's surface structure, which improves the pathogen's capacity for recognition and phagocytosis (97). In this study serum alkaline phosphatase (ALP) activities showed a significant decline in HS and ZNH groups compared to CON and ZN groups. This agreed with Ming et al. (97) and Gulzar et al. (98).