Viral vectors, including lentiviral vectors and adeno-associated viral vectors, have been used as a delivery tool for transduction of neuronal and glial cells with a variety of genetic tools in vitro and in vivo. Although viral vector technologies are essential for application of genetic tools especially in vivo, less attention has been paid to the biological basis of these technologies than to genetic tools delivered. Here we would like to summarize the biological basis of lentiviral vectors and adeno-associated viral vectors and briefly introduce the recent advances from the perspective of the application of these viral vectors to pharmacological research.

As a primate animal model for neuroscience research, the common marmoset (Callithrix jacchus) provides an unprecedented opportunity to gain a better understanding of the human brain function and pathophysiology of neurological and psychiatric disorders, thereby helping in the diagnosis and treatment of those disorders. The marmoset is particularly useful in studying the neural mechanisms underlying social behavior, as their prosocial behavior and visual and vocal communication systems are well-developed. Despite recent advances in biotechnology such as the creation of genetically engineered marmosets, our understanding of the marmoset brain, including its dysfunction in disease, at the circuit level remains limited due to the lack of comprehensive knowledge of the neuronal connections in the marmoset brain. Here we describe the development of genetic and viral engineering techniques for a particular type of neuron in non-transgenic animals. These approaches, combined with rabies viral tracing, imaging, and electrophysiology, will make it possible to map the connectome and relate neuronal connectivity to function in the marmoset brain. Such circuit-level studies will open a new avenue for non-human primate research that can bridge the gap between basic research and human studies.

The development and persistence of drug addiction are associated with the activation and adaptation of the brain reward circuitry, which consists of dopaminergic projection from the ventral tegmental area to the nucleus accumbens (NAc) and the medial prefrontal cortex (mPFC). In cocaine addiction, cocaine-induced activation and neuroplasticity in the brain reward circuitry may contribute to the acquisition and expression of rewarding memory of cocaine, which is critical for the reinstatement of cocaine seeking. However, it remains unclear which neuronal types causally contribute to the retrieval of cocaine-associated rewarding memory. To address this issue, we used DREADD (Designer Receptors Exclusively Activated by Designer Drugs) technology. To selectively suppress mPFC excitatory neurons, we infused an adeno-associated virus (AAV5 or AAV-DJ) vector expressing hM4Di, an inhibitory DREADD, under the control of CaMKII promotor into the mPFC of wildtype mice. To selectively suppress GABAergic neurons, we infused a Cre-dependent AAV (AAV5 or AAV-DJ) vector expressing hM4Di into the mPFC of GAD67-Cre mice or the NAc of vGAT-Cre mice. We found that, in cocaine conditioned place preference paradigm, the activity of mPFC pyramidal and NAc GABAergic neurons is causally related to the retrieval of cocaine-associated memory. The findings suggest that the mPFC-NAc circuit can be a potential therapeutic target for the drug addiction.

Decision-making is a key activity process that influences many aspects of daily living and both mental and physical health. In general, healthy participants reveal rational choice, but patients with neuropsychiatric disorders reveal irrational and risky choice in decision-making. Addiction is one of typical diseases revealed risky decision-making, addicts select risky action and options that confer short-term rewards at the cost of long-term disadvantages. Thus, irrational and risky decision-making is recognized as a core problem in patients with neuropsychiatric disorders, and a better understanding of the mechanisms underlying altered decision-making would provide insights into potential therapeutic approaches for these diseases. However, the neural pathway and substrates underlying these deficits are particularly unknown. Recently, we found that insular cortex is one of key regions for risky decision-making in an animal model of methamphetamine addiction, by using the designer receptor exclusively activated by designer drug (DREADD) technology, and that GABAergic dysfunction in insular cortex is involved in evaluating the subjective value of reward and reward prediction error. These brain dysfunctions would be related to risk taking behavior in addiction. In this review, we introduced the possible neural pathway related to risky decision-making and behavioral changes in choice strategy using adeno associated virus (AAV).

Two third of our body is composed of water molecules. Regulation of water and electrolytes is indeed the most important homeostatic functions. Many diseases, such as heart failure, are associated with disturbance in fluid homeostasis. Surprisingly, water dynamics inside the brain is still largely unknown. In 2012, a new concept referred as “glymphatic system” was proposed by Nedergaard’s group, where aquaporin4 (AQP4) may play an important role as well as sleep. AQP4 is mainly expressed in the central nervous system, especially in the foot processes of astrocytes; surrounding the capillary, beneath pia matter and lining the ventricles. The unique distribution of AQP4 suggest that AQP4 might play a role in brain water homeostasis. The concept of “glymphatic system” is still controversial, and needs to be clarified with new experimental data. This approach will lead to the better understanding of roles of astrocytes in neurodegenerative diseases and pharmacokinetics inside the brain, and eventually will facilitate the development of new drugs for sleep or mental disorders. It has been accumulating evidence that sleep disturbance is related to several kinds of chronic diseases such as hypertension and diabetes. In addition, the number of patients with dementia are significantly increasing. It is therefore critical to understand the physiological and pathological mechanisms of brain lymphatic system from the medical and social point of views. Here I will discuss about the roles of AQP4 in neurodegenerative diseases and introduce new knowledge regarding to “glymphatic system”.

Three-dimensional (3D) cultured hepatocyte capable of maintaining liver-specific function in an in vivo state over a relatively long period of time have drawn attention as a new method for evaluating the metabolic process, hepatotoxicity and enzyme induction potential of drugs. When human hepatocytes were seeded on a plate for spheroid formation, and cell morphology and albumin secretion were examined, hepatocyte spheroid was stably maintained for at least 21 days after seeding. As a result of drug exposure to this spheroid, sequential metabolic reactions by Phase I and Phase II enzymes and metabolic reactions peculiar to only humans were observed. Moreover, when several drugs were exposed to spheroids and hepatotoxicity was evaluated, stable values were obtained for the 50% inhibitory concentration (IC₅₀) of albumin secretion at 14 and 21 days. The IC₅₀ values of most of the tested drugs were lower than in conventional assays, suggesting that the reported evaluation methods might underestimate hepatotoxicity. Furthermore, examination of mRNA expression level and activity of various cytochrome P450 (CYP) after exposure of typical inducers of CYPs to hepatocyte spheroid resulted in a significant increase in the expression level and activity of each. From these results, it was shown that this 3D hepatocyte spheroid system is suitable for follow-up of metabolic processes, long-term tests of hepatotoxicity and enzyme activity induction potential of drugs.

Lenvatinib is an oral multikinase inhibitor that targets VEGF receptors 1–3, FGF receptors 1–4, PDGF receptor α, RET, and KIT. The preclinical studies of lenvatinib for hepatocellular carcinoma (HCC) suggest that lenvatinib exerts the potent antitumor effect on the basis of the inhibitory actions on VEGF and FGF-induced tumor angiogenesis and on FGF-induced tumor cell growth. Phase I and II trials were conducted in Japan and Korea evaluating the maximal tolerated dose, efficacy, and safety of lenvatinib for HCC patients and have produced promising results. Considering the relationship between body weight, AUC and dose in HCC patients, the recommended starting dose was determined to be 8 mg/day for patients weighing lower than 60 kg and 12 mg/day for patients of 60 kg and higher. A phase III REFLECT study have demonstrated that the non-inferiority of lenvatinib to sorafenib in overall survival was confirmed and that lenvatinib was significantly superior to sorafenib in the analysis of progression-free survival and response rate. Based on these results, lenvatinib has been approved for the treatment of patients with unresectable HCC in Japan, US, EU and others this year. Clinical studies of lenvatinib combination therapy with transarterial chemoembolization (TACE) and with immune checkpoint inhibitors are currently on-going. Because of the potent antitumor effect, lenvatinib may change treatment strategy for HCC patients in the future.