Experience-dependent maturation of the glomerular microcircuit

Abstract

Spontaneous and patterned activity, largely attributed to chemical transmission, shape the development of virtually all neural circuits. However, electrical transmission also has an important role in coordinated activity in the brain. In the olfactory bulb, gap junctions between apical dendrites of mitral cells increase excitability and synchronize firing within each glomerulus. We report here that the development of the glomerular microcircuit requires both sensory experience and connexin (Cx)36-mediated gap junctions. Coupling coefficients, which measure electrical coupling between mitral cell dendrites, were high in young mice, but decreased after postnatal day (P)10 because of a maturational increase in membrane conductance. Sensory deprivation, induced by unilateral naris occlusion at birth, slowed the morphological development of mitral cells and arrested the maturational changes in membrane conductance and coupling coefficients. As the coupling coefficients decreased in normal mice, a glutamate-mediated excitatory postsynaptic current (EPSC) between mitral cells emerged by P30. Although mitral-mitral EPSCs were generally unidirectional, they were not present in young adult Cx36(-/-) mice, suggesting that gap junctions are required for the development and/or function of the mature circuit. The experience-dependent transition from electrical transmission to combined chemical and electrical transmission provides a previously unappreciated mechanism that may tune the response properties of the glomerular microcircuit.

Fig. 1.

Sensory deprivation prevents the developmental decrease in coupling coefficients and alters olfactory bulb development. (A) Diagram of whole-cell somatic recordings from pairs of mitral cells that project to the same glomerulus. Hyperpolarizing current injections (range −250 to −400 pA, 0.7 s) into cell 1 (black) elicited a voltage deflection in the cell 2 (red). Each trace represents an average of 7–10 sweeps. Mitral cells at P8 showed higher coupling than at P33. (B) The coupling coefficient (ratio of the hyperpolarization in the test/stimulated cell) decreased after P10. (C) The right olfactory bulb was smaller in this P32 mouse after right naris occlusion at P1. The laminar organization of the olfactory bulb and the projection of apical dendrites to glomeruli were not altered in sensory-deprived animals. The images show slices from contralateral and sensory-deprived bulbs (P > 28), in which Thy1-YFP is expressed in a proportion of mitral cells. (Scale bar, 50 μm.) (D) In young adult mice (P > 28), the coupling coefficient was greater on the side that was sensory deprived. In the example contralateral control bulb shown, a hyperpolarizing current injection (−400 pA, 0.7 s) into cell 1 (black) elicited only a small voltage deflection in cell 2 (red), compared with the larger voltage deflection elicited in the deprived bulb (P < 0.001).

Fig. 2.

Sensory deprivation and genetic deletion of Cx36 prevent developmental changes in membrane conductance. (A) Input resistance decreased in control mitral cells for all time intervals after P7–10 including the contralateral control group (P < 0.001). In contrast, input resistance for the sensory-deprived and Cx36^−/− animals was much higher than the contralateral bulbs or age-matched controls, respectively (P < 0.001, P < 0.002). (B and C) As expected, in Cx36^−/− animals a current pulse caused a hyperpolarization in one cell (black trace), but no electrical coupling in the follower cell (red trace). The confocal image shows 2 biocytin-filled mitral cells from a Cx36^−/− mouse (P34). The margin of the glomerulus is outlined by the dotted line. (Scale bar, 50 μm.)

Fig. 3.

A mitral–mitral EPSC emerges in young adult animals. (A and B) For electrically coupled cell pairs at P7–10, a bAP elicited by a 1-ms current injection (bottom traces) produced a capacitative artifact followed by an inward current (top 2 traces). CBX (100 μM) abolished the NBQX-insensitive inward currents (middle traces), consistent with electrical coupling of the bAP. AP5 (100 μM) and gabazine (5 μM) were included in the extracellular solution. In some cases, NBQX (10 μM) decreased the slow component of the inward current (black cell; Right Top), but had no effect on the peak inward current (red cell; Left). (C) In contrast, for mitral cells at P > 28, short current injection (2.5–3.5 nA, 1 ms) elicited a bAP in cell 1 (red trace, Left Bottom) that produced an EPSC in cell 2 (black trace; Left Top) that was completely blocked by NBQX (10 μM; Left Middle). The EPSC was unidirectional as current injection in cell 2 elicited a bAP (black trace; Right Middle), but did not produce an EPSC in cell 1 (red trace; Right Top). At increased gain (right bottom traces), the cell shows only a capacitative transient. AP5 (100 μM) and gabazine (5 μM) were present in the extracellular solution. (D) NBQX completely blocked mitral–mitral EPSCs in young adult animals (14 of 14 pairs, P < 0.0001). All traces are averages of 7–10 consecutive sweeps.

Fig. 4.

Spontaneous mitral cell activity in sensory-deprived and Cx36^−/− animals. (A) In young adult animals (P > 28) that underwent unilateral naris occlusion on P1, there was surprisingly robust spontaneous activity in the sensory-deprived bulb. The traces show 4 representative 2-s epochs of activity for 1 mitral cell. In contrast, an age-matched mitral cell from a Cx36^−/− (undeprived) animal had very little spontaneous activity. (B) Spontaneous activity was integrated (mV-sec) for mitral cells in the groups of animals shown after determining a baseline during a quiescent period for each cell, before the period of data collection. As discussed in the text, spontaneous activity was actually somewhat greater in the deprived bulb than in age-matched controls, whereas in Cx36^−/− mice, it was reduced to the same degree as by addition of NBQX, AP5, and gabazine (blockers). (C) Mitral–mitral EPSCs were not present in Cx36^−/− mice. Short and long current injections elicited single bAPs or bursts of bAPs, respectively, in either mitral cell (bottom traces), but no synaptic current was present in the corresponding postsynaptic mitral cell (top traces; n = 8). The small response represents capacitative coupling. All traces represent an average of 7–10 consecutive sweeps.