In vivo Intracellular Recording of Neurons in the Supraoptic Nucleus of the Rat Hypothalamus

Intracellular recordings were made from cells in the hypothalamic supraoptic nucleus in the urethane-anaesthetized male rat using the ventral surgical approach. Impalements lasted from 5 min to 1 h and recorded cells had an input resistance of 55 to 170 megohms. Spikes of over 50 mV were recorded from 14 cells which could be antidromically activated by stimulation of the neural stalk. The spikes showed a hyperpolarizing afterpotential and the broadening characteristic of rapidly firing magnocellular neurons, which recovered rapidly (<200 ms). When depolarized, the cells showed evidence of a transient potassium current. Recurrent synaptic coupling between the recorded cell and adjacent cells would be expected to alter the hyperpolarizing afterpotential of an antidromic spike as compared with a spontaneous spike; no perceptible difference in the waveforms of the different types of spike could be detected in 11 spontaneously active cells. Application of just subthreshold stimuli to the neural stalk did not evoke depolarizing or hyperpolarizing potentials. Suprathreshold shocks to the neural stalk, when the antidromic spike was prevented by collision, also had no discernible effect on membrane potential. Thus intracellular recordings from magnocellular neurons in vivo revealed electrophysiological properties similar to those seen in vitro. No evidence for synaptic interconnection between magnocellular neurons was found in male rats.     

Comparison of three intracellular markers for combined electrophysiological, morphological and immunohistochemical analyses.

Hypothalamic paraventricular and supraoptic neurons were recorded intracellularly in coronal slices and injected with Lucifer yellow, ethidium bromide or biocytin. Electrical properties, morphological staining and neurophysin immunohistochemistry were compared among the 3 markers. Lucifer yellow electrodes had a high resistance and frequently blocked during experiments. Neurons recorded with Lucifer yellow electrodes had low input resistances and low-amplitude, broad spikes. Lucifer yellow labeling in whole mount was highly fluorescent, revealing distal dendrites and axons. Of cells injected with Lucifer yellow, 64% were recovered but were faint after immunohistochemical processing. Recordings with ethidium bromide electrodes were similar to controls, although electrode blockage sometimes occurred. Only somata and proximal dendrites of ethidium bromide-filled neurons were visible in whole-mount. Forty percent of cells injected with ethidium bromide were recovered after immunohistochemical processing; these were invariably faint. Recordings with biocytin-filled electrodes were similar to control recordings. Biocytin-filled, HRP-labeled cells showed distal dendrites and often dendritic spines and axons in 50-75-microns sections. Seventy percent of biocytin-injected cells labeled with fluorescent markers were recovered and remained strongly labeled after immunohistochemical processing. Biocytin had the best electrical and staining properties for combined electrophysiological and anatomical studies.