Abstract
Background: Nestin-immunoreactive (nestin-ir) neurons have been identified in the medial septal/diagonal band complex (MS/DBB) of adult rat and human, but the significance of nestin expression in functional neurons is not clear. This study investigated electrophysiological properties and neurochemical phenotypes of nestin-expressing (nestin+) neurons using whole-cell recording combined with single-cell RT-PCR to explore the significance of nestin expression in functional MS/DBB neurons. The retrograde labelling and immunofluorescence were used to investigate the nestin+ neuron related circuit in the septo-hippocampal pathway. Results: The results of single-cell RT-PCR showed that 87.5% (35/40) of nestin+ cells expressed choline acetyltransferase mRNA (ChAT+), only 44.3% (35/79) of ChAT+ cells expressed nestin mRNA. Furthermore, none of the nestin+ cells expressed glutamic acid decarboxylases 67 (GAD(67)) or vesicular glutamate transporters (VGLUT) mRNA. All of the recorded nestin+ cells were excitable and demonstrated slow-firing properties, which were distinctive from those of GAD(67) or VGLUT mRNA-positive neurons. These results show that the MS/DBB cholinergic neurons could be divided into nestin-expressing cholinergic neurons (NEChs) and nestin non-expressing cholinergic neurons (NNChs). Interestingly, NEChs had higher excitability and received stronger spontaneous excitatory synaptic inputs than NNChs. Retrograde labelling combined with choline acetyltransferase and nestin immunofluorescence showed that both of the NEChs and NNChs projected to hippocampus. Conclusions: These results suggest that there are two parallel cholinergic septo-hippocampal pathways that may have different functions. The significance of nestin expressing in functional neurons has been discussed.