Efficient cell-specific and tightly controllable gene expression is highly desirable in studies of gene function and in gene therapy research. The tetracycline-controlled transcription system (Tet-system) was designed for inducible gene expression in mammalian systems (1). However, the original Tet-systems suffered from high levels of leak expression and poor inducibility (2, 3). More recently, a tightly regulated Tet-system has been developed and marketed by ClontechTM. TRE-Tight vectors drive highly inducible and leak proof expression in many cellular systems. However, the limitation of this system is that the induction of gene expression requires a high level of expression of the tetracycline-dependent transactivator which requires the use of strong, typically viral, promoters. Since most cell-specific mammalian promoters are relatively weak inducers of transcription, it is difficult to achieve combined transcriptional targeting and tight Tet-inducible expression. Our aim was to develop a powerful neuron-specific, tight Tet-inducible lentiviral system. We employed a transcriptional amplification (TA) strategy. To express a tetracycline-dependent transactivator (Tet-off) we used a system in which the first copy of the synapsin-1 promoter (SYN) drives expression of a strong chimeric transcription factor Gal4/p65 and modified the second copy of SYN promoter to incorporate a unique set of Gal4 binding sites. In this system Gal4/p65 binds to multiple Gal4 binding sites upstream of the second SYN promoter leading to highly amplified expression of Tet-off. Neuronal specificity, inducibility and levels of gene expression of this system were characterized both in vitro and in vivo. We demonstrate that the TA-amplified SYN-based Tet-regulatable system greatly increased induction of transgene expression in both PC12 cells in vitro (~8-fold compared to non-TA SYN-based system) and hypoglossal motor neurons in vivo (~12-fold) with no loss of neuronal specificity. For injections into the hypoglossal motor nucleus, male Wistar rats (~250-300 g, n = 4) were anaesthetised intramuscularly with a mixture of ketamine (60mg/kg) and medetomidine (250µg/kg). Seven days postinjection, rats were terminally anaesthetised (sodium pentobarbital, 100mg/kg, i.p.) and perfused intracardially. Importantly, this system allowed complete and reversible control of gene expression using doxycycline both in vitro and in vivo. These results will for the first time allow us to deploy TA-based vectors for powerful neuronal-specific expression, which has extremely tight control over its timing, for probing central neural circuits regulating arterial blood pressure.