Effects of NGF and BDNF on baseline glutamate and dopamine release in the hippocampal formation of the adult rat

Abstract

It has been shown using in vitro techniques that BDNF and NGF evoke neurotransmitter release in the hippocampus but this phenomenon has not been demonstrated in vivo to date. We therefore performed in vivo microdialysis in urethane-anesthetized Fischer 344 rats. The microdialysis probe was implanted stereotaxically into the CA1 area of the hippocampus. Three hours after the implantation of the probe, glutamate (Glu) and dopamine (DA) levels had reached a stable baseline. Four baseline samples were collected every 15 min at a flow rate of 1 microL/min. The growth factors were delivered (1 microL/10 min) using a microinjector attached to the microdialysis probe. We found that BDNF and NGF, when administered into the hippocampus, evoked dopamine and glutamate release in a dose-dependent fashion. NGF produced a biphasic response in the release of Glu, and a uniphasic response in the release of DA, both of which were calcium dependent. The neurotransmitter release induced by NGF was blocked by tetrodotoxin, indicating neuronal origin of this response. The BDNF induced release of DA and Glu was decreased in low calcium conditions, indicating that it is at least partially calcium dependent. Furthermore, BDNF-induced neurotransmitter release was partially blocked by pre-treatment with K252a, an antagonist for tyrosine kinase receptors, indicating that BDNF is acting through Trk receptors to induce neurotransmitter release. These results demonstrate a close relationship between the growth factors BDNF and NGF and the neurotransmitters DA and Glu in the hippocampus of intact animals.

Fig. 1. Effects of BDNF on GLU neurotransmission

A = time course for microdialysis results on GLU levels when BDNF was administered via reverse dialysis. X-axis depicts time in min and Y-axis depicts GLU levels in micromolar (μM). BDNF elicited a dose-dependent elevation in GLU levels, with the highest dose (0.3 μg) eliciting the highest response. The BDNF-induced peak started approximately 15 min after the onset of BDNF administration, and lasted for 45–60 min. TTX, low calcium and the TrkB blocker K252a gave rise to significant inhibition of the signal. B = Bargraph illustrating the area under the curve (AUC) analysis of the GLU elevation. BDNF (0.3 μg) gave rise to significantly higher GLU levels than all other conditions (p < 0.05) (indicated by *).

Fig. 2. Effects of BDNF on DA neurotransmission

A = Time course for BDNF effects on DA levels in the CA1. The onset and magnitude of this response was similar to the GLU response to BDNF (see Fig. 1A), with approximately 200% elevation of basal DA levels, beginning 15 min after BDNF and lasting approximately 45 min. B = Area under the Curve (AUC) analysis bargraph. BDNF elicited a significant increase in DA levels, with the highest dose (0.3 μg) giving rise to the greatest increase (indicated by *). Reverse dialysis of TTX gave rise to a reduced and also delayed response, and low calcium as well as the TrkB inhibitor K252a both significantly decreased DA levels, suggesting a TrkB- and calcium-dependent response.

Fig. 3. Effects of NGF on GLU neurotransmission

A = Time course for alterations of GLU levels in response to NGF administered via reverse dialysis. NGF administration gave rise to a bi-phasic GLU response, with the first and smaller peak occurring at 15 min after the onset of NGF administration, and the second and largest peak occurring at 60 min post-injection. The second peak represented a more than 300% increase in GLU levels, which was the strongest response observed in the present study. Low calcium and TTX both ablated both of the peaks, suggesting a calcium-dependent neuronal response. B = Area under the Curve (AUC) analysis of the NGF response. AUC analysis revealed a dose-dependent relationship for NGF effects on GLU, with 0.3 μg BDNF giving rise to the highest GLU levels (p < 0.05) (indicated by *) and the lowest dose not significantly different from the baseline levels (ns). In addition, TTX and low calcium were not significantly different from the baseline levels, suggesting a complete block of the NGF effects.

Fig. 4. Effects of NGF on DA neurotransmission

A = Time course for NGF effects on DA levels in the hippocampus CA1. The magnitude and time course for the response were both similar to those observed in DA levels following a BDNF administration (see Fig. 2A). Contrary to what was observed with NGF administration on GLU levels (Fig. 3A), the DA response to NGF was not bi-phasic and lasted until about 50 min after the onset of NGF administration. B = Area under the Curve (AUC) analysis of DA levels. NGF gave rise to a significant elevation of DA levels, with the highest dose (0.3 μg NGF) representing the highest increase (p < 0.05) (indicated by *). There was a dose–response relationship also for DA levels after NGF administration, and the response to the lower doses of NGF were much greater than what was observed with BDNF administration (Fig. 2B). Reverse dialysis of TTX or low calcium both blocked this effect, and neither TTX nor low calcium yielded levels that were different from baseline levels, indicating a partial block of NGF effects.