TrkB receptor ligands promote activity-dependent inhibitory synaptogenesis - PubMed (original) (raw)

TrkB receptor ligands promote activity-dependent inhibitory synaptogenesis

F J Seil et al. J Neurosci. 2000.

Abstract

Organotypic cerebellar cultures derived from newborn mice were simultaneously exposed to activity-blocking agents and neurotrophins for 2 weeks. Activity-blocked explants treated with the TrkB receptor ligands BDNF and neurotrophin-4 (NT-4) developed a full complement of Purkinje cell inhibitory axosomatic synapses, as defined ultrastructurally, and displayed control spontaneous cortical discharge rates after recovery from activity blockade. Otherwise untreated activity-blocked cultures and activity-blocked cultures exposed to the TrkC receptor ligand NT-3 had reduced inhibitory synapse development and persistent cortical hyperactivity after recovery. The added TrkB receptor ligands did not induce axonal sprouting to account for increased inhibitory synaptogenesis. Addition of neurotrophins to untreated cerebellar cultures did not increase the complement of Purkinje cell axosomatic synapses. Exposure of cerebellar cultures to a combination of antibodies to BDNF and NT-4 resulted in reduced inhibitory synapse formation, similar to the effects of activity blockade, indicating the necessity for endogenous neurotrophins for development of the full complement of inhibitory synapses in the presence of neuronal activity. Application of antibodies to BDNF and NT-4 to cerebellar explants exposed to picrotoxin to increase neuronal activity prevented the hyperinnervation of Purkinje cell somata by inhibitory terminals characteristic of cultures exposed to picrotoxin alone. These results are consistent with the concept that TrkB receptor ligands promote inhibitory synaptogenesis. The ability of neurotrophins to substitute for neuronal activity in encouraging development of inhibitory synapses may have therapeutic implications.

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Figures

Fig. 1.

Fig. 1.

Electron micrographs of Purkinje cell soma profiles from control, activity-blocked, and neurotrophin-treated activity-blocked newborn mouse-derived organotypic cerebellar cultures at 15 DIV. All cells are shown at the same magnification (3150×). The_arrows_ indicate axosomatic synapse profiles. Astrocytic sheaths are evident as relatively clear areas around the circumferences of the cells. A, Purkinje cell from a control culture maintained in standard nutrient medium. B, Purkinje cell from a cerebellar explant continuously exposed to activity-blocking agents, 10−8

m

TTX and 11.1 m

m

Mg2+, incorporated into the nutrient medium. C, Purkinje neuron from a cerebellar culture exposed since explantation to nutrient medium with incorporated activity-blocking agents and 25 ng/ml NT-3. D, Purkinje cell from a cerebellar explant continuously exposed to activity-blocking agents and 25 ng/ml NT-4. There are no significant differences in the morphology of the cells other than the ratios of axosomatic synapse to soma profiles, which are reduced in activity-blocked cultures and activity-blocked cultures treated with NT-3, and at control levels in activity-blocked explants treated with TrkB receptor ligands (Table 1).

Fig. 2.

Fig. 2.

Whole-mount preparations of cerebellar cultures reacted at 15 DIV with antibody to nonphosphorylated neurofilament protein and processed by the peroxidase-antiperoxidase method (375× magnification). Purkinje cell recurrent axon collaterals are indicated by arrows. A, Purkinje cells from a culture continuously exposed to activity-blocking agents.B, Purkinje cells from an activity-blocked culture treated since explantation with BDNF, 25 ng/ml nutrient medium. Axonal sprouting was not evident in neurotrophin-treated activity-blocked cultures.

Fig. 3.

Fig. 3.

Extracellular records of spontaneous cortical activity in cerebellar cultures after 2 weeks in vitro.A, Untreated control explant recorded at 14 DIV.B, Increased activity with spike bursts in a cerebellar explant 40 min after recovery from activity blockade induced by continuous exposure to 10−8

m

TTX and 11.1 m

m

Mg2+. The culture had been silent during the first 10 min after transfer to a medium without activity-blocking agents, in contrast to control cultures, which were immediately active after transfer to a physiological recording medium. Recorded at 15 DIV. C, Cortical spikes recorded 45 min after transfer to a recording medium. The culture had been exposed to a combination of activity-blocking agents and BDNF (25 ng/ml medium) for 15 DIV before recording. The spontaneous cortical activity pattern is similar to that of untreated control cultures. _Time bar_at the bottom, 5 sec.

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