Designer drugs are psychoactive substances that are synthesized by altering the chemical structure of substances of abuse to resemble their pharmacodynamic and pharmacokinetic actions.  Designer drugs exert their effects by affecting the neurotransmitters, endocrine, and/or exocrine systems.  Central (CNS) and peripheral nervous system (PNS) pathologies are usually accompanied by an imbalance in the content of two or more neurotransmitters in specific or multiple regions of the brain.  Designer drugs can impede the neurotransmitter content by disturbing the synthesis, storage, metabolism, release, reuptake, and pre-and/or post-synaptic receptor action (s).  Thus, Designer drugs can be damaging to humans through enhancing or diminishing neurotransmission. Piperazine designer drug’s illegal manufacturing and abuse have been increasing significantly worldwide. In the current study, we investigate the site(s) and mechanism(s) of action of the piperazine designer drugs affecting the cholinergic, and dopaminergic neurotransmission.  At the present time, there are few studies with regard to the hippocampal and dopaminergic neurotoxicity of piperazine derivatives 3-trifluoromethylphenylpiperazine (3-TFMPP), 2-trifluoromethylphenylpiperazine (2-TFMPP) and 4-trifluoromethylphenylpiperazine (4-TFMPP).  N27 rat dopaminergic cells have been used to study the dopaminergic neurotoxic mechanisms of several substances. Furthermore, HT-22 mouse hippocampal cells have been used to study the cholinergic/glutamatergic neurotransmission as well as the etiopathology of several types of dementia.  In this study, we established the mechanisms of dopaminergic and hippocampal neurotoxicity of TFMPP derivatives. TFMPP derivatives caused neurotoxicity in a dose-dependent and time-dependent manner along with the instigation of dopaminergic and hippocampal neuronal cell death.  Moreover, TFMPP derivatives reduced the synthesis and intensified the breakdown of dopamine and acetylcholine significantly which resulted in decreased dopaminergic and cholinergic neuronal viability.  Additionally, TFMPP derivatives prompted oxidative stress, mitochondrial dysfunction, inflammation, as well as apoptosis.  Designer drug manufacture and usage should be firmly controlled to protect society from various CNS and PNS disorders.