Robert C. Gesteland

Comments on Microelectrodes

Metal-filled microelectrodes are best for high-frequency work; fluid-filled ones are best for low frequencies and dc. Both have advantages and drawbacks. This paper gives the results of experience with both sorts of probe. Practical hints and recipes are included because these seldom appear in detail.

Development of olfactory receptor neuron selectivity in the rat fetus

Olfactory receptor neurons begin to differentiate from stem cells on day E10 of embryonic life in the rat. By day E16, the receptor epithelium is well populated. On this day single neuron action potentials could be recorded with some ease and the electro-olfactogram was well developed. The receptor neurons were functional in that they responded to the vapors of odorous substances. However, they were not selective. Each cell responded to nearly all of the substances in the stimulus set. The first synaptic connections between receptors and mitral cells are established on day E18. The olfactory marker protein is reported to appear first in the receptors on the same day. By day E21, single unit responses changed dramatically. The cells became selective, responding to about half of the substances in our set. The electro-olfactogram reached its limiting amplitude well before this time.

Postnatal proliferation and maturation of olfactory bulb neurons in the rat

Mitral cells are formed prenatally whereas most granule cells originate postnatally. Material was taken from 2-day-old, 14-day-old, 28-day-old, and adult rat olfactory bulbs and processed for rapid Golgi or Cresyl Violet staining. We show that the number of granule cell bodies/mitral cell body increases from 7.0 to 46.3 during the first two weeks of life; most mitral cells appear morphologically functional during the first postnatal week; few granule cells appear to be functional until the second postnatal week; and the number of short axon interneurons increases dramatically during the second postnatal week. We conclude the newborn rats have an intact afferent pathway from olfactory receptors to primary cortex that lacks the extensive interneuronal circuitry characteristic of adults.

Assessment of the Sniff Magnitude Test as a clinical test of olfactory function

Olfactory threshold and odor identification tasks currently serve as the standard approaches to the clinical assessment of olfactory function. Although these methods can be used effectively with the average patient, they suffer from some limitations when used to evaluate children, people with cognitive impairment, or people from diverse cultural backgrounds. A novel approach to the clinical evaluation of olfactory function, the Sniff Magnitude Test, attempts to minimize the cognitive demands of an olfactory test and thereby overcome some of the limitations of alternative tests. This is achieved by measuring the reflex-like reduction in sniffing that occurs when a malodor is encountered. The reliability and the validity of the Sniff Magnitude Test were assessed by testing people on two occasions using the University of Pennsylvania Smell Identification Test (UPSIT), a butanol threshold task, the Alcohol Sniff Test, and the Sniff Magnitude Test. The test-retest reliability of the Sniff Magnitude Test was r=.80, higher than the butanol thresholds and Alcohol Sniff Test but somewhat lower than the UPSIT. Performance on the Sniff Magnitude Test (the sniff magnitude to a malodor relative to nonodorized air) was correlated between r=-.61 and r=-.66 with the other measures of olfactory function. This range of correlations was comparable to that observed between the butanol threshold, the UPSIT, and the Alcohol Sniff Test. Finally, evidence for the advantages of the Sniff Magnitude Test, as compared to the UPSIT, was provided by a study with young children. It is concluded that the Sniff Magnitude Test has significant potential as a clinical measure of olfactory function, and that further testing and development of this method are warranted.

Transduction physiology of olfactory receptor cilia

Electro-olfactograms (EOGs) evoked by 8 odorants from frog olfactory epithelia during ciliary regrowth and during epithelial regeneration were analyzed. During ciliary regrowth following detergent-induced ciliary removal, EOG amplitudes initially increase proportionately with ciliary length. EOGs reach maximal amplitudes after 2 days of growth, when cilia are 40 micron long. Therefore olfactory transduction sites are located primarily on cilia rather than on the dendrite terminal and most of the receptor current enters through the proximal portion of the cilium. Zinc sulfate lavage of the nasal cavity causes selective necrosis of the receptor epithelium. During epithelial regeneration, EOGs increase linearly with time from 13 days after zinc lavage, the time of first cilium emergence, through 30 days. The rate of increase is different for different odorants. At 30 days and within a period of a few days, EOG amplitudes increase abruptly, then asymptote. Thus the development of receptors for different substances occurs at different rates and occurs in two steps. The transition between the two developmental states is coincident with arrival of receptor axon terminals at the central nervous system and with the immobilization of the ciliary contractile apparatus. Since there is continual generation of new receptor neurons throughout life, EOGs recorded in a normal nose reflect a complex combination of the differing receptor processes of cells of differing developmental stages.

Speculations on smell.

Experimental research into olfactive code, noting psychological and physiological analysis, suggesting specific chemically sensitive molecules role

Cell and explant culture of olfactory chemoreceptor cells

An in vitro system for the study of maturation of rat and chick embryonic olfactory receptor cells is presented. A variety of dissociating agents, culture media and substrata were tried in attempts to obtain a preparation of mature living olfactory receptor cells readily visible in the microscope. Maturation was judged by the development of axons greater than 1 mm long, by the presence of cilia at the end of the dendrites and, in the rat, by the presence of immunohistochemically demonstrable olfactory marker protein, a protein present only in olfactory receptor cells. By these criteria, dissociated cells did not mature in vitro, though occasional bipolar cells with relatively short axons were seen. In explant culture, small fragments of rat tissue were positive for all 3 criteria after 6 days. In 9-day cultures, the axons had grown up to 3 mm long in both rat and chick cultures. Olfactory bulb fragments co-cultured close to the olfactory epithelium had no influence on the direction of outgrowth of axons from the olfactory receptor cells. Preliminary experiments with intracellular electrodes on the fragment cultures suggest that there are two cell types in the epithelium; one with a potential of -25 to -30 mV and, the other, -12 to -15 mV.

Assessment of the influence of cognition and cognitive processing speed on three tests of olfaction

The extent to which measures of working memory, cognitive speed, and verbal retrieval are associated with performance on tests of olfaction was evaluated in a sample of 138 older adults. Structural equation modeling techniques indicated that verbal retrieval difficulties significantly affect performance on the University of Pennsylvania Smell Identification Test (UPSIT). Further, poor working memory and slow cognitive speed significantly affect performance on the UPSIT and the phenyl ethyl alcohol threshold test. The Sniff Magnitude Test was not influenced by any of the cognitive variables. Odor threshold and identification tasks may overestimate olfactory loss when cognitive impairment is not taken into account.

Transmembrane Currents in Frog Olfactory Cilia

SummaryWe have measured transmembrane currents in intact single cilia from frog olfactory receptor neurons. A single cilium on a neuron was sucked into a patch pipette, and a high-resistance seal was formed near the base of the cilium. Action potentials could be induced by applying suction or a voltage ramp to the ciliary membrane. A transient current was seen in some cells on stimulation with odorants. After excision from the cell, most of the cilia showed increased conductance in a bath containing cAMP, indicating that the cytoplasmic face of the ciliary membrane was accessible to the bath. The estimated resistance of a single cilium was surprisingly low.

A comparison of the sniff magnitude test and the University of Pennsylvania Smell Identification Test in children and nonnative English speakers.

The sniff magnitude test (SMT) is a reliable and rapid clinical test of olfactory function that is minimally dependent on cognitive and linguistic abilities. In this study, we compared performance on the SMT and University of Pennsylvania Smell Identification Test (UPSIT) in samples of children and nonnative English speakers. Previous research has shown that these populations perform poorly on the UPSIT as compared with young, healthy U.S. adults. Such performance differences may reflect variations in memory/cognition and language/culture rather than olfactory abilities. The UPSIT scores of children and of Indian and Chinese graduate students were found to be lower than those of young U.S. adults. By contrast, these groups did not perform more poorly than U.S. adults did on the SMT. The results are consistent with findings from our studies, with the elderly showing that performance on the UPSIT, but not the SMT, is significantly correlated with measures of memory, language and other cognitive abilities. The findings highlight the utility of the SMT when evaluating the olfactory ability of the very young, older adults and people with diverse linguistic and cultural backgrounds.